Advances in Metallodrugs: Preparation and Applications in Medicinal Chemistry [1 ed.] 1119640423, 9781119640424

Table of contents :
Cover
Advances in Metallodrugs:
Preparation and Applications
in Medicinal Chemistry
Copyright
Contents
Preface
1 Metallodrugs in Medicine: Present,
Past, and Future Prospects
2 Chemotherapeutic Potential of
Ruthenium Metal Complexes
Incorporating Schiff Bases
3 Role of Metallodrugs in Medicinal
Inorganic Chemistry
4 Ferrocene-Based Metallodrugs
5 Recent Advances in Cobalt Derived
Complexes as Potential Therapeutic Agents
6 NO-, CO-, and H2S-Based
Metallopharmaceuticals
7 Platinum Complexes in Medicine
and in the Treatment of Cancer
8 Recent Advances in Gold Complexes
as Anticancer Agents
9 Recent Developments in Small
Molecular HIV-1 and Hepatitis B
Virus RNase H Inhibitors
10 The Role of Metals and Metallodrugs
in the Modulation of Angiogenesis
11 Metal-Based Cellulose:
An Attractive Approach Towards
Biomedicine Applications
12
Index

Citation preview

Advances in Metallodrugs

Scrivener Publishing 100 Cummings Center, Suite 541J Beverly, MA 01915-6106

Emerging Trends in Medicinal and Pharmaceutical Chemistry Series Editor: Shahid-ul-Islam and B.S. Butola The Emerging Trends in Medicinal Chemistry and Pharmacology Series is intended to provide recent trends, the state -of-the-art, and advancements particularly in the rapidly growing fields of drug design and synthesis, medicinal natural products, phytochemistry, pharmacology and applications. With a focus on generating means to combat different human diseases, the series addresses novel strategies and advanced methodology to circumvent the invasion from microbial infections and to ameliorate the effects caused by dreadful diseases. Each volume from the series will provide high-level research books covering theoretical and experimental approaches of medicinal natural products, antimicrobial drugs, chemotherapeutic agents, anticancer agents, phytochemistry and pharmacology. The volumes will be written by international scientists for a broad readership researchers and students in biology, chemistry, biochemistry, medicinal science, chemical and biomedical engineering.

Publishers at Scrivener Martin Scrivener ([email protected] Phillip Carmical ([email protected])

Advances in Metallodrugs Preparation and Applications in Medicinal Chemistry

Edited by

Shahid-ul-Islam, Athar Adil Hashmi and Salman Ahmad Khan

This edition first published 2020 by John Wiley & Sons, Inc., 111 River Street, Hoboken, NJ 07030, USA and Scrivener Publishing LLC, 100 Cummings Center, Suite 541J, Beverly, MA 01915, USA © 2020 Scrivener Publishing LLC For more information about Scrivener publications please visit www.scrivenerpublishing.com. All rights reserved. No part of this publication may be reproduced, stored in a retrieval system, or transmitted, in any form or by any means, electronic, mechanical, photocopying, recording, or otherwise, except as permitted by law. Advice on how to obtain permission to reuse material from this title is available at http://www.wiley.com/go/permissions. Wiley Global Headquarters 111 River Street, Hoboken, NJ 07030, USA For details of our global editorial offices, customer services, and more information about Wiley products visit us at www.wiley.com. Limit of Liability/Disclaimer of Warranty While the publisher and authors have used their best efforts in preparing this work, they make no rep­ resentations or warranties with respect to the accuracy or completeness of the contents of this work and specifically disclaim all warranties, including without limitation any implied warranties of merchant-­ ability or fitness for a particular purpose. No warranty may be created or extended by sales representa­ tives, written sales materials, or promotional statements for this work. The fact that an organization, website, or product is referred to in this work as a citation and/or potential source of further informa­ tion does not mean that the publisher and authors endorse the information or services the organiza­ tion, website, or product may provide or recommendations it may make. This work is sold with the understanding that the publisher is not engaged in rendering professional services. The advice and strategies contained herein may not be suitable for your situation. You should consult with a specialist where appropriate. Neither the publisher nor authors shall be liable for any loss of profit or any other commercial damages, including but not limited to special, incidental, consequential, or other damages. Further, readers should be aware that websites listed in this work may have changed or disappeared between when this work was written and when it is read. Library of Congress Cataloging-in-Publication Data ISBN 978-1-119-64042-4 Cover image: Pixabay.com Cover design by Russell Richardson Set in size of 11pt and Minion Pro by Manila Typesetting Company, Makati, Philippines Printed in the USA 10 9 8 7 6 5 4 3 2 1

Contents Preface xiii 1 Metallodrugs in Medicine: Present, Past, and Future Prospects Imtiyaz Yousuf and Masrat Bashir 1.1 Introduction 1.2 Therapeutic Metallodrugs 1.2.1 Anticancer Metallodrugs 1.2.1.1 Mechanism of Anticancer Action 1.2.2 Antimicrobial and Antiviral Metallodrugs 1.2.2.1 Antimicrobial Metallodrugs 1.2.2.2 Antiviral Metallodrugs 1.2.3 Radiopharmaceuticals and Radiodiagnostic Metallodrugs 1.2.4 Anti-Diabetic Metallodrugs 1.2.5 Catalytic Metallodrugs 1.3 Future Prospects 1.4 Conclusion References 2 Chemotherapeutic Potential of Ruthenium Metal Complexes Incorporating Schiff Bases Manzoor Ahmad Malik, Parveez Gull, Ovas Ahmad Dar, Mohmmad Younus Wani, Md Ikbal Ahmed Talukdar and Athar Adil Hashmi 2.1 Introduction 2.2 Schiff Base Complexes of Ruthenium as Anticancer Agents 2.3 Conclusion References 3 Role of Metallodrugs in Medicinal Inorganic Chemistry Manish Kumar, Gyanendra Kumar, Arun Kant and Dhanraj T. Masram 3.1 Introduction

1 2 6 6 7 15 15 16 17 19 22 23 25 26 41

42 43 63 64 71 72 v

vi  Contents 74 3.2 Platinum Anticancer Drugs 3.2.1 Nucleophilic Displacement Reactions in Complexes 80 of Platinum 3.2.2 Mode of the Interaction of Cisplatin Species With Nitrogen Donors of DNA Strand 80 3.2.3 Systemic Toxicity of Cisplatin 82 3.3 Copper-Based Anticancer Complexes 82 82 3.3.1 Copper is Essential for Health and Nutrition 3.3.2 Healthcare Applications of Copper 83 3.3.3 Copper and Human Health Disorders 83 3.3.3.1 Wilson’s Disease (WD) 84 3.3.3.2 Menkes’ Disease 85 3.3.4 Role of Copper Complexes as Potential Therapeutic Agents 85 3.3.4.1 Thiosemicarbazones-Based Complexes 86 3.3.4.2 Quinolones-Based Copper Complexes 88 3.3.4.3 Naphthoquinones 88 89 3.4 Zinc Anticancer Complexes 3.4.1 Biologically Importance of Zinc 90 3.4.2 Schiff Base Chemistry 92 3.4.2.1 Schiff Base and Their Metal Complexes 92 3.4.3 Zinc-Based Complexes 93 94 3.4.4 Top Food Sources of Zinc 3.4.5 Role of Zinc in Human Body 97 3.4.6 Zinc as a Health Benefit 98 3.4.7 Zinc in Alloy and Composites 100 100 3.4.8 Zinc Supplementation as a Treatment 3.4.8.1 Zinc Deficiency 101 102 3.4.8.2 Zinc Toxicity 3.4.8.3 Zinc and Viral Infections 102 3.4.9 Gastrointestinal Effects 103 3.5 Future Prospects of Metallodrugs 103 References 104 4 Ferrocene-Based Metallodrugs Hamza Shoukat, Ataf Ali Altaf and Amin Badshah 4.1 Introduction 4.2 Ferrocene-Based Antimalarial Agents 4.2.1 Mechanism of Action 4.3 Ferrocene-Based Antibacterial and Antifungal Drugs 4.3.1 Schiff Base Derived Ferrocene Conjugates as Antibacterial Agents

115 115 117 118 118 119

Contents  vii 4.3.2 Ferrocenyl Guanidines as Antibacterial and Antifungal Agents 121 4.3.3 Sedaxicene as Antifungal Agents 122 4.4 Ferrocene-Based Anti-Tumor and Anti-Cancerous Drugs 123 4.4.1 Ferricenium Salts as Anti-Tumor Agents 124 4.4.2 Ferrocenylalkylazoles Active Anti-Tumor Drugs 124 4.4.3 Ferrocene Conjugated to Peptides for Lung Cancer 125 4.4.4 Ferrocenylalkyl Nucleobases Potential Anti-Cancerous Drugs 126 4.4.5 Ferrocenyl Sub-Ordinates of Illudin-M 126 4.4.6 Ferrocenyl Derivatives of Retinoids Potential Anti-Tumor Drug 127 4.4.7 Targeting Breast Cancer With Selective Ferrocene-Based Estrogen Receptor Modulators (SERM) 128 4.5 Conclusion 131 4.6 Future of Ferrocene-Based Drugs 131 References 132 5 Recent Advances in Cobalt Derived Complexes as Potential Therapeutic Agents Manzoor Ahmad Malik, Ovas Ahmad Dar and Athar Adil Hashmi 5.1 Introduction 5.2 Cobalt Complexes as Potential Therapeutic Agents 5.3 Conclusion References

137 137 138 153 154

157 6 NO-, CO-, and H2S-Based Metallopharmaceuticals R. C. Maurya and J. M. Mir 6.1 Introduction 158 6.2 Signaling Molecules: Concept of “Gasotransmitter” 160 6.2.1 Therapeutic Applications of NO, CO, and H2S 162 6.2.1.1 Exogenous NO Donating Molecules 163 6.3 NO Donors Incorporated in Polymeric Matrices 167 6.3.1 Metal Nitrosyl Complexes 168 6.3.1.1 Sodium Nitroprusside (SNP) 168 6.4 Dinitrosyl Iron Thiol Complexes (DNICs) 170 6.5 Photoactive Transition Metal Nitrosyls as NO Donors 170 6.6 Exogenous CO Donating Molecules 173 176 6.7 H2S Donating Compounds 176 6.7.1 H2S Gas: A Fast Delivering Compound

viii  Contents 6.7.2 Sulfide Salts: Fast Delivering H2S Compounds 6.7.3 Synthetic Moieties 6.7.3.1 Slow-Delivering H2S Compounds 6.7.3.2 H2S-Releasing Composite Compounds 6.7.4 Naturally Occurring Plant Derived Compounds 6.7.4.1 Garlic 6.7.4.2 Broccoli and Other Cruciferous Vegetables 6.8 Concluding Remarks and Future Outlook References

177 178 178 179 182 182 184 185 186

7 Platinum Complexes in Medicine and in the Treatment of Cancer 203 Rakesh Kumar Ameta and Parth Malik 7.1 What is Cancer? 203 7.1.1 Characteristic Features of Cancer Cells 205 7.1.2 Definition of Anticancer Compound 206 207 7.1.3 Anticancer Attributes of Pt Complexes 7.1.4 Native State Behavior of Pt Complexes 208 7.2 Compatibility of Pt Compounds in Cancer Treatment 209 7.2.1 Significance of DNA as Primary Target 209 7.2.2 Kinetics of DNA Binding Activities 210 210 7.2.3 Structural and Regioselectivity of DNA Adducts 7.2.4 Studies on Action Mechanism 211 7.3 Pt Complexes as Anticancer Drugs 214 7.3.1 DNA-Coordinating Pt(II) Complexes 214 7.3.2 DNA-Covalently Binding Pt(II) Complexes 219 222 7.3.3 Targeted Pt(II) Complexes 224 7.3.4 Pt(IV) Prodrugs 7.3.5 Multiple Action of Pt(IV) Prodrugs 225 7.3.6 Targeted Pt(IV) Prodrugs 228 7.3.7 Photodynamic Killing of Cancer Cell 231 by Pt Complexes 231 7.4 Conclusion Acknowledgments 232 References 232 8 Recent Advances in Gold Complexes as Anticancer Agents Mohammad Nadeem Lone, Zubaid-ul-khazir, Ghulam Nabi Yatoo, Javid A. Banday and Irshad A. Wani 8.1 Introduction 8.2 Evolution of Metal Complexes as Anticancer Agents

247

248 250

Contents  ix 251 8.3 Gold Complexes 8.3.1 Complexes with Nitrogen Donar Ligands 252 8.3.2 Complexes with Sulphur Donar Ligands 254 8.3.3 Complexes with Phosphorus Donar Ligands 255 8.3.4 Complexes with Sulphur-Phosphorus Donar Ligands 256 8.3.5 Organometallic Gold Complexes 259 8.3.6 Miscellaneous 260 262 8.4 Nano-Formulations of Gold Complexes 8.5 Future Challenges and Perspectives 263 8.6 Conclusion 265 Acknowledgements 266 References 266 9 Recent Developments in Small Molecular HIV-1 273 and Hepatitis B Virus RNase H Inhibitors Fenju Wei, Dongwei Kang, Luis Menéndez-Arias, Xinyong Liu and Peng Zhan 273 9.1 Introduction 9.1.1 Activity and Function of HIV and HBV RNases H 274 9.1.2 The Metal-Chelating RNase H Active Site 274 9.2 RNase H Inhibitors and Strategies in the Discovery of Active Compounds 276 276 9.2.1 High-Throughput Screening 9.2.2 Design Based on Pharmacophore Models 278 9.2.3 Novel Inhibitors Obtained by Using “Click Chemistry” 279 9.2.4 Dual-Target Inhibitors Against HIV-1 Integrase (IN) 280 and RNase H 9.2.5 Inhibitors Obtained by Using Privileged Fragment-Based Libraries 282 9.2.6 RNase H Inhibitors in Natural Products 283 9.2.7 Drug Repurposing Based on Privileged Structures 284 9.3 Conclusion 286 References 287 10 The Role of Metals and Metallodrugs in the Modulation of Angiogenesis 293 Mehmet Varol and Tuğba Ören Varol 294 10.1 Introduction 10.2 Metallodrugs in Anticancer Therapy 297 10.3 Angiogenesis as a Substantial Target 300 of Tumorigenesis

x  Contents 10.4 Metals and Metallodrugs in Angiogenesis 10.5 Concluding Remarks and Future Prospects References 11 Metal-Based Cellulose: An Attractive Approach Towards Biomedicine Applications Kulsoom Koser and Athar Adil Hashmi 11.1 Introduction 11.2 History of Cellulose 11.3 The Properties and Structure of Cellulose 11.4 Modification of Cellulose 11.4.1 Acid Hydrolysis 11.4.2 Oxidation 11.4.3 Esterification 11.4.4 Amidation 11.4.5 Carbamiation 11.4.6 Etherification 11.4.7 Nucleophilic Substitution 11.4.8 Further Modification 11.5 Present and Future Medical Applications of Cellulose as Well as Its Components 11.5.1 Cellulose Used as Wound Dressing 11.5.2 Dental Applications 11.5.3 Engineering 11.5.4 Controllable Drug Delivery System 11.5.5 Blood Purification 11.5.6 Wrapping Purpose 11.5.7 Renal Failure 11.6 Conclusion References 12 Multifunctional Nanomedicine Nobel Tomar, Maroof A. Hashmi and Athar Adil Hashmi 12.1 Introduction 12.2 Diagnostics and Imaging 12.3 Drug Delivery and Therapy 12.3.1 Drug Delivery by Organic Nanomaterials 12.3.1.1 Liposomal Drug Delivery 12.3.1.2 Polymeric Drug Delivery 12.3.1.3 Proteins and Peptides for Drug Delivery

302 306 306 319 320 320 321 322 322 324 326 331 333 336 339 341 344 344 345 346 348 348 350 351 351 352 363 364 366 369 369 369 371 373

Contents  xi 374 12.3.2 Drug Delivery by Inorganic Nanomaterials 12.3.2.1 Metal and Metal Oxides 374 12.3.2.2 Au NPs 375 12.3.2.3 Carbon-Based NPs 375 12.3.2.4 Silicon-Based Nanostructures for Drug Delivery 378 12.3.3 Photo Therapy 379 380 12.3.3.1 Photodynamic Therapy 12.3.3.2 Photothermal Therapy 381 12.3.4 Radiation Therapy 383 12.3.5 Neutron Capture Therapy 384 12.4 Regenerative Medicine 385 386 12.5 Future Prospects and Conclusion References 387

Index 403

Preface Over the past few decades, medicinal inorganic chemistry as an interdisciplinary sub-area of bioinorganic chemistry has received the growing attention of researchers in the search for promising antimicrobial, antimalarial, antiviral, and antitumor chemotherapeutic agents. An excellent compilation of reports on metal complexes has revealed the potency of metal complexes as better therapeutic agents. Metal-containing drugs have several promising advantages over organic ligands and have gained the trust of researches after the worldwide approval of the drug cisplatin. Their distinct mechanism of action makes them perfect candidates as alternatives to the conventional drugs to which resistance has already been shown. In this direction, a huge number of transition metal complexes have been synthesized and evaluated for their biological profiles. This book is organized into 12 important chapters that focus on the progress made by metal-based drugs as anticancer, antibacterial, anti­ viral,  anti-inflammatory, and anti-neurodegenerative agents, as well as highlights the application areas of newly discovered metallodrugs. It can prove beneficial for researchers, investigators, and scientists whose work involves inorganic and coordination chemistry, medical science, pharmacy, biotechnology, and biomedical engineering. We are indebted to all the authors for their commitment and for bringing their knowledge and professional experience to making this project a reality. Last but not least, the editors would like to thank Mr. Martin Scrivener, President of Scrivener Publishing, USA, who accepted and supported this project. Shahid-ul-Islam Athar Adil Hashmi Salman Ahmad Khan April 2020

xiii

1 Metallodrugs in Medicine: Present, Past, and Future Prospects Imtiyaz Yousuf* and Masrat Bashir Department of Chemistry, Aligarh Muslim University, Aligarh, India

Abstract

Metal coordination complexes on account of their unique properties of metals which includes variable oxidation states, geometry, coordination numbers, redox behavior, and ability to bind to a wide variety of types of ligands offer a versatile platform for the design of novel therapeutic and diagnostic agents. The therapeutic potential of metal ions can be optimized by tethering it to a suitable frame work that not only tune but synchronize the organic ligand scaffold to act in concord at the target site. Medicinal inorganic chemistry is a growing interdisciplinary field of pharmaceutical research which involves design of therapeutic and diagnostic agents with emphasis on medicinal use for the treatment of various chronic diseases. The serendipitous discovery of cisplatin, inorganic anticancer drug opened up new prospects in the area of medicinal inorganic chemistry that not only cured cancer but provided a continuous spur towards the development of new metallodrugs that can address the serious challenges in the drug regime. Thus, many metal-based therapeutics and diagnostic agents have been explored extensively for their diverse applications as artificial metalloenzymes, DNA foot-printing agents, and nucleic acid structural probes, etc. Given the premises of metallodrugs in the medicinal field, this chapter focuses on the progress made by metal-based drugs as anticancer, anti-bacterial, anti-viral, anti-inflammatory, and antineurodegenerative agents, as well as emphasis on the new strategies to be used in the development of new potential metallodrugs. Keywords:  Medicinal inorganic chemistry, metallodrugs, metal-coordination complexes, therapeutic and diagnostic agents, chronic diseases, drug delivery, prodrugs

*Corresponding author: [email protected] Shahid-ul-Islam, Athar Adil Hashmi and Salman Ahmad Khan (eds.) Advances in Metallodrugs: Preparation and Applications in Medicinal Chemistry, (1–40) © 2020 Scrivener Publishing LLC

1

2  Advances in Metallodrugs

1.1 Introduction Medicinal inorganic chemistry is an interdisciplinary sub-area of bioinorganic chemistry field which tethers the applications of inorganic chemistry and biological disciplines, thereby investigate the intriguing properties of metal ions, their complexes, and other metal binding compounds for the therapeutic and diagnostic purposes [1–6]. Conceptually, the field of medicinal inorganic chemistry includes the biomimetic chemistry of metal ions in metalloproteins [7, 8], identification of metal ions in pathogenic protein misfolding [9, 10], functions of endogenous and exogenous metal ions at the molecular level [11, 12], and the homeostasis of metal ions in living systems [13]. The use of several metals (Cu, Au, Ag, Hg, and As) can be traced back to ancient civilizations (Mesopotamia, Egypt, India, and China) [14] with the recognition by the Egyptians who used copper to sterilize water with an understanding of disinfection and the Chinese and Arabs utilized gold in the treatment of many chronic diseases [15]. Zinc was found to promote healing of wounds while mercurous chloride was used as a diuretic. Paul Ehrlich the “founder of chemotherapy” developed arsenical, Salvarsan, as a drug for the treatment of Syphilis in early twentieth century (Figure 1.1) [16]. Thus, a link between the discovery of a new elements and their application into the medicinal armamentarium (therapeutic and diagnostic) has been exploited since antiquity (Table 1.1). Numerous metal ions and their complexes have been routinely administered to patients for therapeutic and diagnostic benefit such as platinum and ruthenium complexes in cancer therapy [17–20], gold complexes as anti-arthritis agents [21, 22], cobalt complexes as antiviral [23], and gadolinium and technetium as magnetic resonance imaging (MRI) contrasting agents [24–26] (Figure 1.2). Metal ions can serve many important functions in the biological systems; (i) functional role, i.e., the biological activity is due to direct binding OH

OH NH2

H2N HO

As As As (a)

NH2

HO

OH As As As As As

H2N NH2 OH

H 2N HO

NH2 NH2

(b)

OH

Figure 1.1  Structures of arsenic-based therapeutic drug, salvarsan (3‒amino‒4‒ hydroxyphenyl‒arsenic(III) compounds).

Metallodrugs in Medicine  3 Table 1.1  The use of metal salts and their compounds as therapeutic and diagnostic agents. Metal

Metal-based salt/compound

Therapeutic/diagnostic use

Ag

Silver sulphadiazine

Antibacterial

Al

Al(OH)3

Antacid

As

Salvarsan, Melarsen, Tryparsamide

Antimicrobial

Au

Gold(I) thiolates Auranofin

Antitumour Antiarthritis

Ba

Barium sulphate

X‒ray contrast

Bi

Bismuth subsalicylate, colloidal bismuth citrate, ranitidine bismuth citrate

Antacid, antiulcer

Cu

Copper histidine complex Casiopeinas

Menkes disease Anticancer

Co

Coenzyme B1 Doxovir

Supplement Antiviral

Fe

Sodium nitroprusside Fe(III) desferrioxamine chelates

Vasodilator Antimicrobial

Gd

Gd metallotexaphyrin (Magnevist, Dotarem)

MRI contrast agent Radiopharmaceuticals

Hg

Mereurochrome

Antiseptic

Li

Li2CO3

Manic depression

Pt

Cisplatin, carboplatin, oxaliplatin, nedapltin etc

Anticancer

Ru

NAMI‒A, KP10109, RAPTA‒C etc

Anticancer

Sb

Pentostam, N‒methylglucamine antimonate

Antileishmanial

Tc

99m

Diagnostic imaging

Ti

Titanocene dichloride, bis(β‒ diketonato) Ti(IV)

Anticancer

Tc (V) propyleneamine oxime

(Continued)

4  Advances in Metallodrugs Table 1.1  The use of metal salts and their compounds as therapeutic and diagnostic agents. (Continued) Metal

Metal-based salt/compound

Therapeutic/diagnostic use

V

Bis(maltolato) oxovanodium(IV) Bis(glycinato) oxovanodium (IV) Bis(methylpicolinato oxovanadium (IV)

Antidiabetic

W

PolyoxometalJates

Anti-HIV activity

Zn

ZnO Zn(II)bicyclam complexes Zinc citrate/sulphate

Skin ointment Antiviral Supplement

Zr

Zr(lV) glycinato

Antiperspirant

O

O P

O-

NH3

Cl

N

Pt Cl

O

(a) As

NH2

O H N

N H

HO

H

O O O

O O

H

AcO OH

AcO AcO

O (c)

O

O

(b)

S

O

Gd O

O

O

N

N

O

NH3

O

S

Au

P

OAc (d)

Figure 1.2  Prominent examples of metal-based drug in medicinal inorganic chemistry: (a) Cisplatin, (b) MS–325, (c) Darinaparsin, and (d) Auranofin.

Metallodrugs in Medicine  5 of the metal fragment to the target site [27], (ii) structural role, i.e., the shape of the complex is determined and binding to the biological target occurs through non-covalent interactions [28–30], (iii) act as a carrier for active ligands that are delivered in vivo [31, 32] and protect the ligand before its delivery at the target site, (iv) metal complexes behave as a catalyst in vivo by the production of reactive oxygen species (ROS) that cause cell damage [33, 34], and (v) metal complexes which are photoactive can act as photosensitizers [35, 36]. Metal ions once introduced into a bio system for therapeutic or diagnostic effect can also be removed from back by the judicious use of the chelating ligands (chelation therapy). Many proteins and enzymes bind one or more metal ions to perform their functions where the metal ion is involved in the catalytic mechanism or stabilizes the tertiary and quaternary structure of proteins. Whereas the small organic drug molecules rely purely on carbon, their binding geometry in space is dictated by the hybridization, viz., sp (linear), sp2 (trigonal-planar), and sp3 (tetrahedral) as compared to the diverse geometry in 3D space open to metal-based drugs [37]. Besides linear, square planar, and tetrahedral geometries, pyramidal, trigonal bipyramidal, and octahedral shapes can be created and even higher coordination numbers and geometries with larger metal ions are possible, all of these geometries exhibit a tremendous importance for biological phenomena that allow the fine-tuning of their chemical reactivity in terms of both kinetics (rates of ligand exchange) and thermodynamics (strengths of metal-ligand bonds, redox potentials, etc). Not only the metal but also the ligands can play important roles in biological activity, ranging from outer-sphere recognition of the target site to the activity of any released ligands and ligand centered redox processes. Modification of substituents or ligands around the metal center, thus modulates drug entities to perform manifold functions and at specific target sites to combat chronic diseases, viz., cancers, HIV-AIDS, cardiovascular, cerebrovascular diseases, and respiratory disorders [38]. Since metal ion can participate in biological redox reactions, and many transition metals (Pt, Ru, Fe, Cu) possessing variable oxidation states offer many possibilities for strategic designs of new chemotherapeutics. Many literature reports reveal that the redox properties of the metal ions or the ligands can influence the mechanism of action of metal-based anticancer chemotherapeutic drugs [39–41]. Thus, in metal complexes, it is possible to trigger a desired biological response at the site of action and at the optimum time by controlling the activation process by substitution (ligand exchange) and/or redox processes.

6  Advances in Metallodrugs One of the biggest challenges in enhancing the therapeutic potential of a metallodrug is its delivery to the selective targets. It is imperative for a prospective drug to demonstrate sufficient reactivity towards the selective biological target but less affinity towards other biomolecules encountered on the way which render its deactivation. Prodrugs are drug derivatives that can undergo in vivo transformation to release the active species, with improved physiochemical, biopharmaceutical, and pharmacokinetic properties [42, 43]. The application of prodrug strategy encompasses the use of polymeric conjugate materials and other inclusions of metallodrug in liposomes, protein macromolecules, lipid-based systems, and dendrimers as drug carriers that limit its interaction with biomolecules other than the selected targets [44]. For metal-based therapeutics, this prodrug activation can be accomplished by in vivo ligand substitution, photochemical process and/or redox reactions before reaching the target site. It is thus important to ascertain the active part of the metal complex which is essential for therapeutic activity; the metal itself, the ligands and the intact delivery system. Thus, a rational state-of-art design of therapeutic and diagnostic agents is required to achieve specific targeting features and control toxicity (side effects) which can be achieved by controlling thermodynamic and kinetic processes of metal complexes.

1.2 Therapeutic Metallodrugs 1.2.1 Anticancer Metallodrugs Cancer is a class of disease in which a group of cells display uncontrolled growth (division beyond the normal limits), invasion (intrusion and destruction of adjacent tissues), and sometimes metastasis (spread to other locations in the body via lymph or blood) [45]. Current statistics indicate that one in every three people will develop some form of cancer during their lifetime. It is estimated that by 2030, there will be 21.4 million new cases diagnosed every year [46]. Most cancerous cells divide uncontrollably to form lumps or masses of tissue called tumors but some like leukemia (where cancer prohibits normal blood function by abnormal cell division in the blood stream) do not [47]. The complexity of the disease however, arises mainly due to the fact that cancers evolve from different tissues of origin, shows multiple etiologies or endless combinations of genetic or epigenetic alterations. The primary treatment modalities include surgery, chemotherapy, radiations, and immunotherapy, etc [48]. However, the mainstay treatment is based on chemotherapy which a viable alternative

Metallodrugs in Medicine  7 involving various natural and synthetic origin compounds that can kill or halt the unwanted proliferation of cancerous cells. Metal-based antitumor chemotherapeutics gained prominence after the phenomenal serendipitous discovery of the archetypical inorganic drug, cisplatin (cis–diamminedichloroplatinum(II), [cis–(NH3)2PtCl2]) by B. Rosenberg in 1965 [49]. Cisplatin is one of the most effective chemotherapeutic drug used for treating solid malignancies, viz., bladder, melanoma, non–small cell lung, small cell lung, head and neck, cervical, ovarian, and testicular cancers (>90% cure rate) [50]. Currently, it is used in 32 of 78 treatment regimens in combination with a wide range of other drugs including: topoisomerase II inhibitors (doxorubicin, etoposide, and bleomycin), mustards (cyclophosphamide, melphalan and ifosfamide), and antimetabolites [51].

1.2.1.1 Mechanism of Anticancer Action Cisplatin initially enters inside the cell via both passive diffusion and active uptake where it undergoes a ligand substitution event prior to DNA binding. Noticeably, inside the bloodstream (extracellular), cisplatin is relatively stable and maintains its neutral state, due to the high concentration of chloride ions (∼100 mM). However, inside the cell (intracellular), the relatively low chloride ion concentration (∼4−12 mM) causes cisplatin to undergo aquation, in which a chloride ligand is replaced by a water molecule resulting in the formation of cis‒[Pt(NH3)2Cl(H2O)]+ species having half-life of ca. 2 h for the aquation reaction. The positive charged platinum complex is a potent electrophile that is attract to the negatively charged nuclear DNA at the N7 position of purine bases of DNA with a release of the water molecule [52]. The remaining cis-monochloride species ([Pt(NH3)2Cl(H2O)]+) is then subsequently aquated diaqaua species ([Pt(NH3)2(H2O)2]+) allowing the cisplatin to cross-link to another purine. Moreover, the square-planar geometry of cisplatin facilitates ligand substitution, which is necessary for it to form the DNA lesions that characterize its activity. Cross-linking between adjacent guanine residues is considered to be crucial to the cytotoxicity of cisplatin. Such cross-links can occur between deoxyguanosines on the same strand or on different strands, giving rise to intrastrand and interstrand DNA cross-links, respectively. The 1,2‒d(GpG) intrastrand cross-link is the most prevalent lesion (65%), but 1,2‒(ApG) (25%) and 1,3‒d(GpTpG) (10%) intrastrand cross-links also form along with small amounts of GG interstrand crosslinks [53]. These adducts interfere with cellular DNA replication and transcription processes causing eventual cell cycle arrest and potentially activation of pro-apoptotic signals (Figure 1.3) [54].

8  Advances in Metallodrugs CI

Bloodstream [CI–] = 100 mM

C

Plasma Proteins, Albumin

I H3N Pt CI TR

OCTI-3

NH3

INACTIVATION

Passive Diffusion

O

O HO

H OH H N

O OH

O

INACTIVATION

x B flu 7 Ef ATP e tiv gh Ac rou th

NH2

N H

NH3 Pt NH 3

GS

INACTIVATION

DN Rep A air

CI l + mM oxo 10 H3N Pt OH2 Cyt – ] = 4– n o i t I a NH3 INACTIVATION [C CI Aqu MT H3N Pt CI NH3

Transcription Inhibition

CELL DEATH!

INACTIVATION

Figure 1.3  The anticancer action of cisplatin revealing the extracellular and intracellular events that influence its anticancer activity. Reproduced with permission from ref [54(c)]. Copyright 2013 The American Chemical Society.

Regardless of the therapeutic success of cisplatin in the treatment of several types of tumors, its effectiveness is severely hindered by adverse side effects, viz., alopecia, ototoxicity, neurotoxicity, myelosuppression, and nephrotoxicity [55]. Another major drawback is tumor resistance, either acquired or intrinsic resistance [56]. However, considerable efforts are being made by many research groups around the world to mitigate the severe side effects, provide oral bioavailability, and overcome resistance issues of cisplatin; and consequently, a plethora of second generation platinum analogs were envisaged, viz., carboplatin, oxaliplatin, nedaplatin, heptaplatin, and satraplatin (Figure 1.4) [57]. Carboplatin and oxaliplatin have entered worldwide in chemotherapeutic drug regimens as a first-line treatment for colorectal cancer [58]. Carboplatin is primarily used against ovarian cancers; however, it has also found use in treating a diverse type of cancers including retinoblastomas, neuro- and nephroblastomas, brain tumors, as well as cancers of the head, neck, cervix, testes, breast, lung, and bladder [59]. Carboplatin has the same implications as that of cisplatin, but with a different toxicity profile [60]. Another

Metallodrugs in Medicine  9

H3N

O

O

N H2

O

O

O

O

H3N

Pt

Pt H3N

O

H2 N

O

Pt O

H3N

O

O

Carboplatin

Oxaliplatin

Nedaplatin

O

O

O NH2

O

CI

Pt NH2

O

O

CI

Pt O

NH3

NH2 Pt

N H2

O

NH2

O

Lobaplatin

O O

Satraplatin

O O

Heptaplatin

Figure 1.4  Structure of anticancer platinum metallodrugs; carboplatin and oxaliplatin (approved), Carboplatin, Lobaplatin, heptaplatin, and satraplatin (in clinical trials).

front-line platinum anticancer drug oxaliplatin has gained global approval for combination chemotherapy treatment against colon cancers [61] and was subsequently approved for clinical use in countries like France and the United States [62]. Oxaliplatin features two chelating ligand groups with oxalate and R,R‒diaminocyclohexane (DACH) as leaving and non-leaving groups, respectively. Nedaplatin has been mainly used to treat head, neck, and esophagus cancers besides small cell lung and non‒small cell lung cancers in Japan [63]. The drug possesses cis ammine as non-leaving group along with a chelating leaving group ligand as glycolate, which confers greater water solubility than cisplatin. Heptaplatin was developed in Korea and is being used against gastric cancer under the market name SunPla. The drug contains two types of chelating ligands, a malonate as a leaving group and 2‒(1‒methylethyl)‒1,3‒ dioxolane‒4,5‒dimethanamine as its non-leaving ligand. However, the search for efficacious drugs that overcome the limitations of platinum compounds such as severe side effects, high systemic toxicity, and incidence of drug resistance have motivated researchers to introduce non-platinum drugs into the drug regimens. No-platinum drug entities are likely to have different mechanism of action, bio-distribution, toxicity profile, and could be effective against human cancers that are poor chemosensitive or have become resistant to conventional platinum drugs. Three-dimensional transition metals particularly Ti, Fe, Co, Cu, and Zn have invoked considerable interest as antitumor chemotherapeutics as these metal ions are site selective at physiological pH and are compatible to the biological system in contrast to platinum-based anticancer agents. Although essential metal ion that escapes from its normal metabolic

10  Advances in Metallodrugs pathway could show toxic effects in an organism, complexes of such metals can serve as effective cytotoxic agents. Among first row transition metal ions titanium complexes, viz., titanocenedichloride, (Cp2TiCl2) and budotitane (Figure 1.5) have demonstrated pronounced antitumor properties and low toxic side effects [64]. Cp2TiCl2 which has entered in phase II clinical trial inhibits DNA synthesis rather than RNA and protein synthesis and titanium accumulates in nucleic acid rich regions in tumor cells after in vivo or in vitro administration [65]. The complex also showed weaker affinity to DNA bases and binds more strongly to phosphate backbone. Budotitane, which was the first non-platinum transition metal anticancer agent to be tested in clinical trials, was quite effective against a number of ascites tumors and induced colorectal tumors [66]. Clinical trials indicated that it was fairly well tolerated by patients with the dose limiting side effects being cardiac arrhythmia [67, 68]. Iron is the most significant essential metal ion in biology which serves as important cofactors of many redox enzymes [69]. Iron is vital for wide variety of metabolic processes including oxygen transport, DNA synthesis, and electron transport reactions. Many synthetic iron complexes have been reported displaying anticancer activities that are often linked to the redox reactions of Fe(II) or Fe(III) under physiological conditions [70]. One of notable example of anticancer compounds involving iron complexation is bleomycin which was clinically used to treat testicular carcinoma with high cure rates [71]. Bleomycin is a glycopeptide comprising a N-terminal metal-binding domain that coordinates to Fe(II) ion through five nitrogen donor atoms of amines, pyrimidine, and imidazole [72]. The coordination of dioxygen to Fe(II) is followed by one-electron oxidation which generates a bleomycin-Fe(III) OOH species. This species induced DNA damage as well as production of ROS, leading to apoptotic cell death [73]. Recently,

Cl

Cl

O

Ti O

O C2H5 O C2H5

Ti O

O

Figure 1.5  Structure of Titanium anticancer agents: (a) Titanocene dichloride and (b) Budotitane.

Metallodrugs in Medicine  11 an organometallic compound Ferrocifen, an analog of Tamoxifen (which has been widely used in the clinic for the treatment of hormone dependent breast cancers) was discovered indicating a distinguished mode of antiproliferative activity (Figure 1.6) [74]. Another noteworthy example of iron complexes is a polypyridine iron(II) complex, Fe(II)–N4Py {N4Py = N,N–bis(2–pyridylmethyl)–N–bis(2–pyridyl)methylamine} which is synthetic bleomycin mimetic [75]. The complex was reported to cleave DNA efficiently under aerobic conditions and induced cell death and caused nuclear DNA damage [76]. Copper is one of widely distributed in the biological system and is the most familiar redox metal accessible within the cellular potential range [77]. Due to the plasticity and participation of copper as an integral part of the active site of metalloproteins (superoxide dismutase, ceruloplasmin, cytochrome oxidase, and tyrosinase), it familiarizes its coordination with the human body’s functions [78]. Copper binds to electron rich nucleic acids (DNA/RNA) with higher affinity than any other divalent cation and induces conformational changes in polynucleotides and bio-membranes. The altered metabolism of cancer cells and the differential response between normal and tumor cells to copper is the basis for the development of copper complexes endowed with antitumor properties [79]. Ruiz-Azuara et al. have synthesized a series of Cu(II) complexes with diimine ligand donors having the trade name “CasiopeinasⓇ, (Cas)” as antitumor chemotherapeutics (Figure 1.7) [80, 81]. These compounds are mixed chelate copper(II) complexes with a general condensed formula [Cu(N–N)(A–A)][NO3], where N–N represents neutral diimine donors, either phen or bipy, A–A stands for uninegative N–O or O–O donors, either aminoacidates or acetylacetonate. The activity of Cas II– gly, [Cu(1,4–dimethyl–1,10–phenanthroline)(glycine)NO3], a novel anticancer agent, was tested against two cell lines, L1210 (murine leukemia) 2+

2+

OH

NCCH3

H3CCN R

N N N

Fe

OH

(a)

N

Fe

N N

R HO

N

Fe N

N

N

OH

H3CCN R = H, Ph

(b)

Figure 1.6  Examples of some iron anticancer agents: (a) Ferrocifen (ferrocene derivative of tamoxifene) and (b) Iron(II) pentapyridyl complexes.

12  Advances in Metallodrugs + H3C

CH3

N

N

_ NO3

H3C

CH3

N

Cu H2N

N

+

_ NO3

Cu O

C

O

O

O

Figure 1.7  Structure of Cas II–gly and III–ia.

and CH1 (human ovarian carcinoma). It was observed Cas II–gly was highly active against these cell lines, including cell lines resistant to cisplatin and mechanism of cell death was both via apoptosis and necrosis [82]. Another variant of the “Casiopeina” series, [Cu–(acetylacetonato) (4,4′–dimethyl–2,2′–bipyridine](NO3), (CasIII–ia), was found to exhibit antineoplastic effects on glioma C6 [83]. Ruthenium(II) complexes are rapidly becoming a prime focus for the development of new more efficacious metal-based anticancer drug entities due to their unique spectroscopic and electrochemical properties [84, 85]. Ruthenium is well suited for pharmacological applications as it offers various oxidation states (II, III, and IV) under physiologically conditions [86]. Plethora of ruthenium complexes have been synthesized and have successfully demonstrated significant cytotoxic and antimetastatic properties with reduced side effects [87]. The first successful breakthrough in the area of ruthenium-based chemotherapeutics was achieved by B. K. Keppler et al. who synthesized imidazolium [trans–RuCl4(1H–imidazole)(DMSO–S)] (NAMI–A) and indazolium [trans–RuCl4(1H–indazole)2] (KP1019), as a substitute to platinum-based drugs (Figure 1.8). These drugs have successfully completed phase I clinical trials and are now undergoing further clinical evaluation [88, 89]. Both NAMI–A and KP1019 are ionic Ru(III) compounds bearing structural novelty possessing negatively charged octahedral metal center coordinated to heterocyclic nitrogen donor ligands and equatorial chlorides; a protonated form of the heterocyclic nitrogen ligands as counter ions that are replacable by sodium or other cations [90]. NAMI–A in solution phase involves both loss of Cl− and DMSO. NAMI–A is the most intensively studied ruthenium anticancer complexes because of its ability to prevent the metastasis formation or inhibit the growth of secondary tumor cells while KP1019 is active only against primary cancers [91]. KP1019 has entered in the clinical trials after it demonstrated in vitro cytotoxic activity against

Metallodrugs in Medicine  13

HN H N+

N CI CI

Ru S

O

CI

CI

CI

N H

CI

HN

N Ru

N

NH CI

HN

H N+

CI

CH3 CH3

(a)

(b)

Figure 1.8  Ru(III) anticancer compounds currently in clinical trials: (a) imidazolium [trans-RuCl4(1H–imidazole)(DMSO–S)] (NAMI–A) and (b) indazolium [trans-RuCl4 (1H-indazole)2] (KP1019).

cisplatin-resistant human colon carcinoma cell lines [92] along with efficient in vivo activity against various tumor types. Pertinent to mention, Ru(II) compounds that are administered to the patient are not as the active species rather Ru(III) complexes are first reduced into a more active Ru(II) form (Scheme 1.1). The plausible mechanistic hypothesis for Ru(III) compounds was attributed on “activation by reduction” mechanism, according to which the Ru(III) complexes act as prodrugs that can be reduced to Ru(II) active species in the hypoxic (therefore reducing) environment of cancer cells [93]. The hydrolysis reaction of Ru–X bonds to give ruthenium‒aqua species (aquation) is an important aspect of the therapeutic behavior for ruthenium complexes. The corresponding aqua species OH2 Ru2+

Target site

+H2O –X

Active Species

Ru3+ A prodrug

Ru2+ +H+

Ru3+

–H+

OH

Scheme 1.1  A generalized scheme depicting possible action for Ru(III) prodrugs invoking “activation by reduction” hypothesis (X = Cl−,Br−, I−).

14  Advances in Metallodrugs exist over a wide range of pH, but for pH > pKa, the hydroxo species formed by deprotonation are predominant. Since, the hydroxide is a less labile ligand than water, it will not so easily be displaced by biomolecule targets. “Aquation” of the chloro complexes may be suppressed extracellularly due to high chloride concentrations (0.1 M) but because of lower intracellular Cl‒ concentrations (4–25 mM), the aquation reaction is highly possible. Many ruthenium compounds have been found are non-toxic and have been quite selective for cancer cells. This has been attributed to the ability of ruthenium to mimic iron in binding to biomolecules. As cancer cells overexpress transferrin receptors to satisfy their increased demand for iron, ruthenium-based drugs have been found to be delivered more efficiently to cancer cells [94]. In recent years, half-sandwich–configured organoruthenium(II)–arene scaffold have emerged as a versatile tool for the design novel anticancer agents because their biological activity and pharmacological properties can easily be modulated by ligand selection [95]. The different mode of action is a consequence of their high lipophilicity that favor better cellular uptake; and the presence of labile ligands, viz., chlorido/carboxylato, favor the extracellular binding with the drug target. Besides possessing supposedly low general toxicity and high selectivity of ruthenium-arene complexes towards cancer cells, the big reasons for the flourishing design of arene-ruthenium–based anticancer drugs are the amphiphilic properties of the arene ruthenium unit, which provides the hydrophobic nature to the arene ligand counter balanced by the hydrophilic metal center [96]. The pioneering work of Paul J. Dyson and P. J. Sadler in the field of anticancer organometallics led to successful revelation of two lead Ru(II)‒arene anticancer agents, RAPTA–C, and RAED which are at an advanced preclinical development stage (Figure 1.9) [97, 98]. +

Ru

Cl

P Cl

N (a)

N

H2N N

_ PF6

Ru

Cl

NH2 (b)

Figure 1.9  Structures of anticancer organoruthenium complexes (a) RAPTA–C ([RuII(cym)(PTA)Cl2], PTA = 1,3,5–triaza–7–phosphatricyclo[3.3.1.1]decane; cym = η6–p–cymene) and (b) RAED ([RuII(η6–biphenyl)(en)Cl]+.

Metallodrugs in Medicine  15

1.2.2 Antimicrobial and Antiviral Metallodrugs 1.2.2.1 Antimicrobial Metallodrugs The therapeutic use of metals and their salts like that of copper, zinc, silver, etc., have been used against microbial infections since early civilization [99, 100]. The metals and their compounds have been shown to possess antimicrobial effects primarily via in-cell redox activity and their sporadic interference with native metal cofactors even at low concentration [101]. However, aqua or other simple metal ions owing to their generally low solubility in water, poor stability, and resultant potential toxicity in humans were not found suitable for therapy. Previously used drugs atoxyl, arsanilic acid, against trypanosomiasis (sleeping sickness) diseases provided an upsurge towards the development of metallodrugs against microbial diseases [102]. In this purview, metal complexes based on As, Ag, Au, Sb, and Cu, etc., have shown efficient antimicrobial and antifungal activity [103]. Although the pioneering work of Ehrlich et al. used arsenic-based antimicrobials called salvarsan have been widely used [104]. The therapeutic effectiveness of salvarsan is due to the formation of oxidized arsenoso compound, which is the active form that binds to sulphydryl (−SH) compounds thereby showing the antimicrobial effect [105]. However, other arsenic-based metallodrugs including Melarsoprol is still being used against “African sleeping sickness” disease under the market name “Arsobal” despite its severe toxic issues [106]. Antimony-based complexes Sb(III) tartarmetic and potassium antimony tartrate have also previously been used against different types of leishmaniasis [107]. Other Sb(IV) drugs that are being used include sodium stibogluconate (Pentostam) and melglumine antimoniate (Glucantim or Glucantime) [108]. Silver complexes have found to possess efficient pharmacological benefits from antibacterial activity, anti-inflammatory to difficult-to-heal wounds [109]. Silver-based FDA approved drugs, viz., silver sulphadiazine (SilvadeneⓇ and FlamazineⓇ), have been used in wound therapy while a dilute solution of AgNO3 is being used for the prophylaxis of bacterial conjunctivitis in infants. Cerium nitrate‒silver sulphadiazine (Flammacerium) is being employed for the treatment for most cutaneous burns and to reduce the inflammatory response to burn injury [110]. The pharmacological activity of silver is predominantly attributed to its interference with the electron transport system or cell membranes and partly to its interaction with thiol groups of the vital enzymes [111]. Two promising metallodrug candidates that are currently undergoing phase II clinical trials are the ferrochloroquine (antimalarial agent) and VT1161 (antifungal agent).

16  Advances in Metallodrugs NH2

N

HN

N

H2N

Fe

N N

N

Cl

S S

(a)

(b)

Ag

N

O H2N

OH

As

HN

S O N

N

N

N Ag O

Ag O N

N N

Ag

S O

O NH2 n

(c)

Figure 1.10  Antimicrobial and antiparasitic drugs approved and in clinical trials: (a) ferroquine, (b) melarsoprol, and (c) silver sulphadizine.

In conjugation with ferrocene, the chloroquine (a well-known antimalarial drug) has addressed the resistance issues of malaria pathogens against chloroquine drug (Figure 1.10) [112].

1.2.2.2 Antiviral Metallodrugs In recent times, many metal complexes have been screened for their antiviral activities including that of Cu(II), Co(II), Zn(II), etc., however, very less success has been made so far. To the best of our knowledge, only two metal complexes have successfully proven to be effective against viral diseases which are at their clinical stages [113]. These cobalt(III) metallodrugs called as Doxovir (CTC‒96) has successfully completed phase II clinical trials for the treatment of Herpes virus and phase I clinical trials for the treatment of viral eye infections (adenoviral conjunctivitis) [114]. The biochemical action of Doxovir, [Co(acacen)(L)2]+, was attributed to its possible interaction with proteins by coordinating to histidine (His) residues involving a dissociative exchange pathway for labile axial ligands, L (L = 2‒methylimidazole). Many polyoxometallates which include polynuclear, transition-metal oxyanions (POM), like [NaW2lSb29O86] [NH4]17 and K12H2[P2W12O48].24H2O have demonstrated efficient antiviral activity. The nanoscale assemblies of early

Metallodrugs in Medicine  17 O

+

NH N N O

Co N

N

– + ONa

S Hg

O

N H

(a)

(b)

Figure 1.11  Two antiviral metallodrugs: (a) Doxovir (CTC‒96) and (b) Sodium thiomersal.

transition metals, viz., vanadium, tungsten, molybdenum, etc., with oxygen have resulted in the formation of cage-like structures and bind to positive patches of HIV gp120 blocking binding to lymphocyte CXCR4 receptor (Figure 1.11) [115, 116]. Sodium‒2‒ethylmercurithio‒benzoate, thimerosal aka thiomersal, is mainly added as a preservative. The ethylmercurithio cation of thiomersal binds readily to thiol‒groups in protein structures blocking their enzymatic activity. It has been reported that aluminum and mercury-based compounds were used as adjuvants in vaccines. Aluminum hydroxide, aluminum phosphate, and potassium alum (KAl(SO4)2·12H2O) have been found help to stimulate the immune response [117].

1.2.3 Radiopharmaceuticals and Radiodiagnostic Metallodrugs Radiometals are radioactive isotopic elements that are being harnessed for applications in medical diagnosis and therapeutics. The radioactive property of many radioisotopes have been utilized for diagnostic imaging techniques, viz., 67Ga, 99mTc, 111In, 177Lu,68Ga, 64Cu, 44Sc, 86Y, 89Zr, etc., as well as for radiotherapy applications (47Sc, 114mIn, 177Lu, 90Y, 212/213Bi, 212 Pb, 225Ac, 186/188Re) [118]. However, in order to synergize these isotopes for specific biological applications, the “free” radiometal ions are to be sequestered from aqueous solution by using specific chelating ligands to avoid transchelation and hydrolysis. Interestingly, the structural features of radiometal–chelate complexes have demonstrated significant impact on the overall pharmacokinetic and pharmodynamic properties of a radiopharmaceutical, and noticeably, many radiometal complexes have shown facile renal renal excretion [119]. Recently, many radiodiagnostic pharmaceuticals are being successfully used in cancer diagnosis, cardiological disorders, kidney or liver abnormalities, and many neurological disorders [120]. Technetium‒99m (99mTc) has demonstrated marked dominance in radio diagnostic applications which is reflected by the two leading 99mTc

18  Advances in Metallodrugs heart imaging agents Cardiolite and Myoview [121, 122]. However, key challenge of the 99mTc isotope is its ongoing shortage of production which has sparked the search for possible future alternatives in radiochemistry. MRI is a sophisticated medical diagnostic tool used to provide images of the human body parts along with their detailed features [123]. The image contrast has been found to depend on differences in relaxation times and proton density between different tissues [124]. However, to make a distinction between two adjacent regions which is often hampered by the inherently narrow relaxation rates of water protons is often a challenge. As a result of this, efforts have been directed towards increasing tissue contrast through the administration of small molecules containing paramagnetic metal ions that are capable of improving contrast by shortening the relaxation time of nearby protons. The larger magnetic moment and long electronic relaxation time of Gd(III) ions makes them ideal in the use of MRI contrast agents [125]. However, due to the high toxicity of its free form, the Gd3+ metal ions have been complexed with suitable chelating ligands, viz., diethylenetriaminepentaacetic acid [H5DTPA] and 1,4,7,10‒tetraaza‒ cyclododecane‒1,4,7,10‒ tetracetic acid [H4DOTA] and are successfully as injectable macrocyclic contrast agents for MRI scans (Figure 1.12).

O

_

_ O

O N

_

O

N

O

N

N

O

N Gd3+

O

+ O

N

C

O

O (a)

C

O

N O

Gd3+

O

N

O

_

_ O

N

O

O

C

C N O

O

N

N

(b)

_ O

_ O

C

Tc

_

O

O

N C

O

_

(c)

Figure 1.12  Prominent examples of metal-based diagnostic metallodrugs: (a) [99mTc(sestamibi)]+ (Cardiolite), (b) [Gd(DOTA)]− (Dotarem), and (c) [Gd(DTPA)]2− (Magnevist).

Metallodrugs in Medicine  19 The imaging agents [Gd(DTPA)]2− (Magnevist) and Gd(DOTA)]− (Dotarem, Gadovist) were approved by FDA 1988 and 2013, respectively [126]. Currently, many efforts are being carried out by focusing on the addition of cell binding ligands on the external surfaces and on the determination of the detection limits associated with these complexes within physiological limits.

1.2.4 Anti-Diabetic Metallodrugs Diabetes mellitus is a type of chronic metabolic disease caused due to alteration in glucose homeostasis and is characterized by persistent hyperglycaemia that results in absolute or relative deficiency in insulin secretion from the pancreatic (beta) cells [127]. The disease has also been attributed to the dysfunction of insulin receptors that show some association with diabetes [128]. It has been reported that around 347 million people are suffering from diabetes mellitus worldwide and it is believed that 80% of diabetes deaths occur in low- and middle-income countries [129]. Many organic drugs such as dipeptidyl peptidase IV (DPPIV) inhibitors, sulphonylureas, metformin, and α‒glucosidase inhibitors, etc., have been used for the treatment of this disease but these drugs have shown limited efficacy and are often associated with severe side effects [130]. Potential insulin mimetic metallodrugs based on vanadium salts and coordination compounds have demonstrated various insulin-enhancing and antidiabetic effects [131]. The vanadium complex, bis(maltolato)oxovanadium(IV) (BMOV), and its ethylmaltol analog bis(ethylmaltolato) oxovanadium (IV) (BEOV) have shown increased in vivo bioavailability over vanadyl sulphate and thereby are the most suitable drug candidates for diabetes treatment (Figure 1.13) [132].

O

O

O

O

V O

O (a)

O O

O

O

O

V O

O

O

(b)

Figure 1.13  Potential metallodrug candidates for the treatment of diabetes at clinical stages: (a) bis(maltolato)oxovanadium(IV) (BMOV) and (b) ethylmaltol analog bis(ethylmaltolato) oxovanadium (IV) (BEOV).

20  Advances in Metallodrugs It has been further reported that the insulin enhancing effect of these vanadium complexes stems from the activation of the insulin receptor through the suppression of insulin receptor tyrosine kinase (IRTK) associated phosphatases [133]. The successful intervention of vanadium compounds for the treatment of diabetes have triggered extensive research on vanadium(IV and V) complexes which have been tethered with diverse group of organic ligands, viz., maltol, kojic acid, picolinic acid, acetylacetonate, dicarboxylate esters, etc., to tune its pharmacological properties incorporate functionalities that increase their therapeutic potential. These organic-vanadate complexes especially vanadate esters demonstrate their pharmacological effects by acting as potential phosphatase inhibitors which is revealed by the formation stable vanadium-phosphatase protein structures [134]. In recent years, many other metals including chromium, zinc, and tungsten have been potentially exploited for development of efficient antidiabetic metallodrugs. Chromium in its trivalent state has emerged as an important inorganic cofactor involved in insulin-based metabolic chemistry and plays a prominent role in carbohydrate metabolism by enhancing insulin signalling towards insulin-sensitive tissues [135]. The importance of Cr(III) complexes as antidiabetic has been summarized by recent examples of antidiabetic and their activity (Table 1.2). Table 1.2  A summary of potential antidiabetic CrIII complexes. Potential ligands for CrIII complexes

Activity

Ref.

Nicotinates, phenylalanines, histidinates

Decrease in blood glucose level

[136]

Propionates

Amelioration of insulin resistance

[137]

Picolinates

Improvement of serum lipid metabolism Improvement of glucose metabolism

[138]

Rutin, folate, and stachyose ligands

Control of blood glucose

[139]

Malate

Control of blood glucose level, liver glycogen level, and of the activities of aspartate transaminase, alanine transaminase, and alkaline phosphatase

[140]

Metallodrugs in Medicine  21 Sodium tungstate, Na2WO4 owing to its specific restoration of hepatic glucose metabolism by the stable oxoanion [WO4]2− has been widely investigated. The [WO4]2− ion has also been shown to mimic most of the metabolic effects of insulin and stimulate insulin output [141]. Sodium molybdate Na2MoO4 has been instrumental in preventing or treating of diabetic mellitus in the early stages of the disease [142]. Zinc being an important constituent of insulin has been found to play a profound role in the physiology and functioning of the enzyme [143]. It is coordinated to three nitrogen atoms from histidine residues along with three water molecules in an overall distorted octahedral environment [144]. The safe oral administration of ZnCl2 was found to exert in vivo antidiabetic effects by possibly by stimulating the adipocyte lipogenesis in rats. The effect of an increased dietary zinc intake at the onset of diabetes mellitus (type 1) led to its prevention by blocking NF‒ƙB activation in the pancreas was tested [145]. Many orally active insulin-mimetic zinc complexes have been successfully screened for their antidiabetic activity against experimental diabetic animals (Figure 1.14). These studies have revealed the importance of zinc in inhibiting the various insulin receptors, PI3K, glucose transporter (GLUT), and phosphodiesterase (PDE) which are associated with insulinomimetic and antidiabetic activities [146]. + Cl

_

N

O

H O Zn

O

O H

O

O

O

O

O Zn

O + _N Cl

(a)

O

O (b)

O S

S Zn

N S

N

O

N

S Zn

S

S

O

N

O (c)

(d)

Figure 1.14  Structures of orally active Zn complexes showing potential antidiabetic effect: (a) Zn(car)2, (b) Zn(mal)2, Zn(pdc)2, and Zn(tanm)2, car; carnitin, mal; malto, tanm; S‒allixine‒N‒methyl, pdc; pyrrolidine‒N‒dithiocarbamate.

22  Advances in Metallodrugs

1.2.5 Catalytic Metallodrugs The potential of metal complexes to act as catalysts in living systems has been exploited by various researchers that resulted in the development of a new class of metallodrugs called as “catalytic metallodrugs” [147]. Catalytic metallodrugs essentially mimic the catalytic reactions of metalloenzymes that occur inside biological systems [148]. The presence of both a catalytic metallic center and a targeting domain mediate the localization of each reactive catalyst to the target. Catalytic metallodrugs bind and irreversibly modify the target during a catalytic process resulting in an enzymelike behavior and being highly stable under biological conditions [149]. Apart from having novel mechanisms of action which can circumvent the development of drug resistance, catalytic metallodrugs can potentially be administered in smaller doses and with lower toxicity. In recent years, there has been substantial upsurge in the development of metal complexes which act as catalytic metallodrugs, viz., the mimics of superoxide dismutases (SOD’s) [150], ROS generators [151], artificial proteases and nucleases, thiol oxidants, and transfer hydrogenation catalysts [152]. Moreover, the catalytic metallodrugs have successfully been explored for their C–C bond formation, deprotection and functional group modification, degradation of biomolecules, and redox modulation. Copper(II) and nickel(II) complex of ATCUN ligands (peptide with amino terminal) have been shown to efficiently cleave proteins [153] and RNAs [154]. Amyloids proteins, peptide deformylases, streptavidins, carbonic anhydrases, and other proteins have been successfully undergone cleavage reactions with [CoII(cyclen)] derivatives, CuII‒EDTA‒biotin or CuII‒1,10‒phenanthroline‒arenesulphonamide derivatives (Figure 1.15) [155]. Remarkably, recent examples of metal-based catalytic drugs include Cu(I)/TBTA-type compounds that help in vivo labeling of proteins, Pd(0) nanoparticles activate anti-cancer drugs, Cu(II)‒ATCUN compounds which target and degrade hepatitis C and HIV RNA, and transfer hydrogenation reactions by using Ir(III) and Ru(II) half sandwich complexes [156, 157]. J.C. Joyner and J.A. Cowan have developed a class of catalytic metallodrugs (Figure 1.16) involving chelating ligands, viz., DOTA, EDTA, NTA, etc., which were chemically linked to the lysine side chain of lisinopril with transition metal ions Fe3+, Co2+, Ni2+, and Cu2+. The resulting M‒chelate‒ lisinopril catalysts could bind to sACE‒1 (human angiotensin converting enzyme which is important therapeutic target for the treatment of hypertension and heart failure) irreversibly inactivate sACE‒1 with high selectivity [158]. Furthermore, it was observed that the oxidative nature of these

Metallodrugs in Medicine  23 O N H

N H H N

NH

M

HN

M NH

NH

M-Cyclam

N

HN

O

O

M-Cyclen

O N

M

O

O

O

O

M-DOTA

O O _ O O

N

O N

M

N

O O

N O

M-DPTA

O

N O

O

M

O

O O

O

O O

N N H

M

_ O

N N N H

M-NTA

M-GGH

Figure 1.15  Transition metal chelates that show efficient catalytic activity, M = Fe3+, Co2+, Ni2+, Cu2+.

M‒chelate‒lisinopril catalysts distinguish them from the relatively slow hydrolytic catalysts while lisinopril. The authors further evaluated many chelating compounds for their “HIV‒1 RRE mRNA” activity in order to corelate the inactivation of the hepatitis C virus by the catalytic metallo­ drugs [159].

1.3 Future Prospects The recent achievements in the development of metal-based pharmaceuticals have stimulated significant progress in the field of medicinal inorganic chemistry which leads to design and synthesize some prominent metal-based diagnostic or therapeutic agents. This rapid growth stems from the spectacular successes of the most notable drug cisplatin. Recent interesting developments in the area of medicinal inorganic chemistry feature in the imaging research, which involve the exploitation of nuclear and electronic properties of the metal ions in vivo, by fluorescence, electronic properties, positron emission, or gamma emission. Moreover, a large variety of physiologically stable gold(III) complexes have been developed that possess significant in vitro and in vivo anti-cancer activities. Several classes

24  Advances in Metallodrugs O O

O

O

O O

N

M

N

O

HN

O

HN HO

N

O O

M

O

O

O HO

N

N H

O

N

N H

O

HO

O

O

M-NTA-lisinopril

HO

O

M-EDTA-lisinopril O O

N

HN

HO

N M

N

N H2

O

N H

O N

N H O

HO

M-DOTA-lisinopril O

Figure 1.16  Potential metal‒chelate‒lisinopril catalysts; M = Fe3+, Co2+, Ni2+, Cu2+.

of anti-cancer platinum(II) complexes which could overcome cisplatinresistance have also been successfully developed and screened against many cancer cell lines. The successful application of bleomycin and NAMIrelated compounds for anti-cancer treatments has inspired researchers to take up iron(II) and ruthenium(II) anticancer complexes for various therapies. Many metal-based complexes with anti-viral effects based on ruthenium and a vanadium have exhibited significant anti-HIV-1 activities. Despite tremendous advancement in the field of metallodrugs, treatment options are still limited for many types of diseases including cancers. The incidences of indiscriminant “off-target” side effects in addition to lack of selectivity, poor biodistribution, and drug resistance limit the effective use of existing conventional chemotherapeutic regimens. Therefore,

Metallodrugs in Medicine  25 identification of new drug design and development of therapeutic strategies that could target diseased cells leaving normal cells unaffected and show high efficiency without inducing any resistance phenomena still continues to be a challenge. Much recently, drug delivery systems have been developed to prevent the shortcomings of conventional metal-based therapy and with a prominent increase in its efficacy. Such systems offer a promising solution by encapsulating metal-based drugs, thereby providing a protective housing for the drug that limits its interaction with healthy cells. More specifically, nanoparticle-based drug delivery systems have emerged as important treatment protocol to improve cancer therapies due to their capacity for overcoming biological barriers and to modulate payloads intracellular trafficking. Nano-sized colloidal carriers of drugs can be regarded as an advanced development in pharmacotherapy. Recent reports have successfully highlighted a comprehensive analysis of various platinum-polymer complexes, micelles, dendrimers, liposomes, and other nanoparticles for the delivery of platinum-based drugs for improving drug efficacy, reducing unwanted side effects and circumventing drug resistance [160]. The nanometer size ranges can significantly enhance the drug delivery by affecting the bio-distribution and pharmacodynamics of metallodrugs.

1.4 Conclusion Medicinal inorganic chemistry is a growing interdisciplinary field of pharmaceutical research which involves design of therapeutic and diagnostic agents with emphasis on medicinal use for the treatment of various chronic diseases. Since many empirical evidences regarding the therapeutic use of metal salts and their compounds has been reported since antiquity but the serendipitous discovery of inorganic anticancer drug, cisplatin opened up new prospects in the area of medicinal inorganic chemistry that not only cured solid cancers but provided an impetus towards the development of new innovative strategies to address the challenges of variety of diseased states. Consequently, a number of metallodrugs are being screened for their use in modern medicine in the treatment and diagnosis of a different diseased conditions related to cancers, infections, HIV-AIDS, diabetes, cardiovascular, inflammatory, and the related syndromes. Metal-based complexes offer a great potential for therapeutic and diagnostic purposes owing to their unique electronic and spectroscopic properties that allow diverse coordination number and geometries apart from their favorable redox potentials accessible at the physiological conditions. The metal itself

26  Advances in Metallodrugs serves as structural center for organizing the organic ligands in the biologically relevant chemical space while the ligand framework plays significant role in metal-based pharmaceuticals via alteration in the biological properties by modifying reactivity or substitution inertness. The choice of metal ions is also a critical feature for the design of metal-based chemotherapeutic drugs. In this chapter, we have focused on the diagnostic and therapeutic metallodrugs which are either in clinical stages or have been FDA approved. Although a significant number of metal-based drug entities are in clinical trials that are being examined both for therapy and diagnosis, but very few have been approved globally. There is still an urgent need for the discovery of new metallodrugs with novel mechanism of action particularly for the diseased conditions that develop resistances towards the current drug regime. The contemporary research on metallodrugs demands the development of therapeutic agents that can mimic the roles of natural metalloenzymes in therapies, hence to develop catalytically active metal drugs. In conclusion, it can be inferred that metallodrugs hold a tremendous potential to help mankind to overcome drug resistance and to find new cures in medicine.

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2 Chemotherapeutic Potential of Ruthenium Metal Complexes Incorporating Schiff Bases Manzoor Ahmad Malik1*, Parveez Gull1, Ovas Ahmad Dar1, Mohmmad Younus Wani2, Md Ikbal Ahmed Talukdar1 and Athar Adil Hashmi1† 1

Bioinorganic Laboratory, Department of Chemistry, Jamia Millia Islamia, New Delhi, India 2 Chemistry Department, Faculty of Sciences, University of Jeddah, Jeddah, Kingdom of Saudi Arabia

Abstract

Cancer is an unfortunate rapid and uncontrolled division of cells which costs life. Different chemotherapeutic drugs and treatment strategies have been employed to combat this dreadful disease. Chemotherapy can be used alone or in combination with other treatments. Over the last couple of years, many advances have been made to curb the uncontrolled division of cells or completely eradicate the cancer. Metal-based drugs lead the combat against this disease owing to their enhanced therapeutic activities and relatively less side effects. Apart from the successful Platinum-based complexes, Ruthenium complexes have been actively studied as metallodrugs for cancer therapy. They have engrossed special attention of medicinal chemists due to their unique properties like variable oxidation states and their ability to mimic iron in their mode of action. They also display selective antimetastatic properties with reduced systemic toxicity. Schiff base complexes of Ruthenium have been recognized to possess broad range of applications and are gaining considerable interest in the area of anticancer drug research. We opine that Ruthenium complexes can prove as apt alternatives for the frequently used cisplatin in the near future. The present chapter compiles examples of some Ruthenium complexes of Schiff bases possessing promising anticancer chemotherapeutic potential. *Corresponding author: [email protected] † Corresponding author: [email protected] Shahid-ul-Islam, Athar Adil Hashmi and Salman Ahmad Khan (eds.) Advances in Metallodrugs: Preparation and Applications in Medicinal Chemistry, (41–70) © 2020 Scrivener Publishing LLC

41

42  Advances in Metallodrugs Keywords:  Schiff bases, Ruthenium complexes, anticancer, cytotoxicity

2.1 Introduction Medicinal inorganic chemistry is receiving growing attention of researchers in the search for promising antibacterial, antifungal, antimalarial, antiviral, and antitumor chemotherapeutic agents [1, 2]. Excellent compilation of reports on metal complexes has unveiled the potency of metal complexes as better therapeutic agents compared to organic molecules [3]. To overcome the shortcomings, organic compounds containing hard and soft donor sites are used as ligands and chelated with transition metals to increase their biological profile [4]. This stimulated more and more researchers to drive their efforts towards this field and develop more potential drugs to combat diseases of current concern. Schiff bases as a class of versatile ligands are increasingly used due to their promising biological applications [5]. Metallation of these versatile ligands with both essential and non-essential elements render them as able candidates to treat various diseases [6]. Huge reports on Schiff base complexes have discussed their antimicrobial, antiviral, and anticancer potential [7, 8]. Among the diseases of present concern, cancer is the second leading cause for increasing number of deaths worldwide. Undoubtedly cisplatin is the most appreciated anticancer drug in the world but suffers from severe side effects like anaemia, diarrhoea, alopecia, petechiae, emetogenesis, fatigue nephrotoxicity, ototoxicity, and neurotoxicity [9]. Facing such undesirable side effects, attempts are being made to develop non-platinum–based metal complexes which are suitable alternatives to cisplatin [9]. Recently, several ruthenium complexes have emerged as successful candidates against this horrible disease [11–16]. Ruthenium metal belonging to the platinum group (ruthenium, rhodium, palladium, osmium, iridium, and platinum) has some distinctive characteristics like variable oxidation states and mimics iron in its mode of action, thus enabling it to bind with proteins such as human serum albumin protein and transferrin protein [17]. Ruthenium complexes have a few similarities with cisplatin-type complexes and their activity mostly depends on the nature of ligand environment, coordination geometry and their versatile redox properties. Ruthenium chelates generally exhibit less toxicity to non-malignant cells and tissues and also evoke drug resistance to a lesser extent as compared to platinum complexes [18–20]. Two prominently known ruthenium complexes KP1019 (trans[RuCl4(Ind)2]IndH; Ind = indazole) and NAMI-A (trans-[RuCl4(DMSO)

Chemotherapeutic Potential of Ruthenium  43

HN Cl Cl

H N N Ru N

Cl

N

NH

Cl NH

N

Cl

Cl

Ru

Cl H3C

S

Cl

H N N H

O

CH3 KP1019

NAMI-A

Figure 2.1  The structure of KP1019 and NAMI-A.

(Im)]ImH; Im = imidazole) (Figure 2.1) have entered clinical trials and showed promising results [21, 22]. It is believed that the ruthenium metal chelates exert their action by forming adducts with specific targets like DNA, proteins, and enzymes but the exact mechanism is still to be explored. In the present chapter, some advances in the field of ruthenium-based antitumor drugs are briefly highlighted with a special emphasis on Schiff base ruthenium complexes.

2.2 Schiff Base Complexes of Ruthenium as Anticancer Agents Cancer is a growing concern affecting people of all ages all over the world. Several strategies are adopted by researchers to treat this global menace [23]. Cisplatin has proved as a model for the development of new anti­ cancer drugs and heralded a new era in cancer treatment [24]. Cisplatin and its analogs (Figure 2.2) are highly effective chemotherapeutic agents and some of them have received clinical approvals [25]. A plethora of other transition metal complexes have been designed and assessed for their anticancer potential and Ruthenium complexes have stood out as potential alternatives to mimic cisplatin in its properties [22, 26]. Combination of biologically active ligands to minimize toxicity toward normal cells is a feasible way to manipulate ruthenium complexes to improve their anti-­ proliferative activity [27]. Such ligands present diverse coordination modes, more compatibility with the biological environment and encourage higher cellular uptake. Overall, the vast potential and versatility of ruthenium complexes confirm the importance they adopt in medicinal chemistry

44  Advances in Metallodrugs H3N Cl Pt H3N Cl Cisplatin

H2 O N Pt N O H2

O

O H3N Pt H3N O

O

O

H3N

Oxaliplatin

Carboplatin

Pt

H3N

O

O

Nedaplatin O

O H2 O N Pt O N H2

O O

O

Lobaplatin

H2 O N Pt O N H2

Heptaplatin

O

O

O

H3 O N Pt N O H3

O O

O O

Dicycloplatin

O

Figure 2.2  Molecular structures of some platinum anticancer drugs that are approved or undergoing clinical trials.

and surely certify their increasing emergence in the combat against cancer [28]. A number of ruthenium complexes displaying anticancer activity are reported in the literature, justifying their importance. Schiff base compounds being endowed with potential biological applications are engaged to chelate with metal atoms to enhance their biological profile [29]. Several Schiff base complexes have displayed promising anticancer activity [8, 30]. Here, we will discuss the Schiff base complexes of ruthenium as anticancer agents. Two different ruthenium complexes (1.0 and 1.1) (Figure 2.3) were separated from a one pot reaction of an equimolar amount of salicylaldehydethiosemicarbazone and [RuHCl(CO)(PPh3)3]. The newly synthesized complexes were characterized by various spectroscopic techniques and it was interesting to note that in one of the complexes 1.0, the ligand coordinated through NS (nitrogen and thiolate sulfur) acting as a bidentate ligand forming a four-membered ring and in complex 1.1, it coordinated through ONS donor atoms. Both the complexes were screened for their comparative biological studies like DNA binding, cytotoxicity (MTT, LDH,

NH2

H

O N

N

N S Ph3P Ru PPh3 OC Cl 1.0

O Ph3P

N

NH2

S PPh3 CO

Ru

1.1

Figure 2.3  New Ruthenium(II) complexes with cytotoxic effects comparable to that of cisplatin.

Chemotherapeutic Potential of Ruthenium  45 and NO), and cellular uptake with an attempt to compare their modes of chelation with their potential biological activities. The IC50 values of complexes 1.0 and 1.1 on the proliferation of human lung cancer (A549) and liver cancer cell lines (HepG2) were found to be 26 μM ± 1.03 and 23 μM ± 0.99 for A549 and 23 μM ± 1.01 and 20 μM ± 1.21 for HepG2 cells, respectively. In comparison with the commonly used standard cisplatin, complex 1.1 was found to have a lower IC50 value for the A549 cell line. However, for HepG2, both complexes have IC50 values higher than that for cisplatin. Biological results established that complex 1.1 displayed higher activity than complex 1.1, that could be due to the strong chelation and increased delocalization of electrons favoring the lipophilic character of metal ions into the cells [31]. Ruthenium complexes (1.3 and 1.4) (Figure 2.4) with N,O–donating heterocyclic hydrazones were synthesized and screened for their in vitro cytotoxicities. The complexes were evaluated for their inhibitory effects on HeLa and MCF-7 cell lines by MTT assay. The IC50 values of the complexes are shown in the Table 2.1 which is indicative of the fact that both the complexes possess cytotoxicity against both the cell lines. Among the two complexes, complex 1.4 showed more promising results than complex 1.3 which may be due the presence of heterocyclic ring containing sulfur in it [32]. Three new mononuclear ruthenium complexes (1.5–1.7) (Figure  2.5) were synthesized and characterized by spectroscopic and analytical

N N O

H

N

O O

Ru PPh 3 Ph3P OC

Ru

Ph3P

1.3

S

NH PPh3

O 1.4

Figure 2.4  Molecular structure of biologically active Schiff base ruthenium(II) complexes.

Table 2.1  IC50 (μM) concentration of ruthenium(II) hydrazone complexes. Complex

HeLa

MCF-7

1.3

18.0 ± 0.5

2±1

1.4

21.0 ± 0.6

6±1

Cisplatin

53.0 ± 0.7

10 ± 2

46  Advances in Metallodrugs H

Cl

Ru N N

PF6

OCH3

Cl Ru N

1.5

PF6

H

Cl

N

1.6

PF6

Ru N NH N 1.7

Figure 2.5  Molecular structure of mononuclear ruthenium(II) complexes showing moderate cytotoxic effects against MCF7, HepG2, Caco-2, and HepG2 cancer cell lines.

techniques. Single crystal X-ray studies confirmed a pseudo-octahedral three-legged, piano-stool geometry around Ru(II), with the ligand coordinated to the ruthenium(II) through two N atoms. All the complexes were screened for their cytotoxic effects against three human cancer cell lines and were tested for their selectivity against non-cancerous human epithelial kidney (HEK 293) cells. The compounds displayed selectivity toward the tumor cells in contrast to the known anti-cancer drug 5-fluoro uracil (IC50< 50). All the compounds exhibited moderate activity (IC50 > 50–100) against MCF7 (human breast adenocarcinoma), but exhibited low antiproliferative activity (IC50 > 100) against Caco-2 and HepG2. Also, the complexes were studied for their antimicrobial activities against six Grampositive and four Gram-negative bacteria [33]. Three tetradentate (ONNO) 1.8–2.0 (Figure 2.6) symmetric and asymmetric Schiff base ruthenium(III) complexes were synthesized and studied for their in vitro anticancer studies using three cancer cell lines: human renal cancer cell (TK10), human melanoma cancer cell (UACC62), and human breast cancer cell (MCF7) using the SRB assay with parthenolide as a standard. Antioxidant activities revealed that the ruthenium(III) complexes showed strong scavenging activities against 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2′-azinobis-3-ethylbenzothiazoline-6-sulfonic

H3C

N O

Cl N

CH3

H3C

Ru

O OH2

HO 1.8

•2H O 2

OH

N

Cl N

O

O OH2

CH3

Ru

HO 1.9

H3C

Ru

•2H O 2

CH3

N Cl N O

O OH2

CH3 •2H O 2

HO 2.0

Figure 2.6  Proposed structure for the Ru(III) complexes 1.8–2.0 investigated for their in vitro anticancer activity.

Chemotherapeutic Potential of Ruthenium  47 acid (ABTS) radicals. The IC50 (90 ± 5- > 100 µM) values obtained suggested that the complexes have low-to-moderate in vitro antiproliferative effect compared to parthenolide (1.43–11.3700 µM) [34]. A series of ruthenium thiosemicarbazone complexes (Figure 2.7) were synthesized by microwave-assisted method. The general formula of the complexes is [(diimine)2Ru(TSC)](PF6)2 where TSC is bidentate thiosemicarbazone ligand and the diimine is either 2,2-bipyridine or 1,10-phenanthroline. The complexes showed promising anticancer activity against human colon cancer cells, Caco-2 and HCT-116. According to obtained results, the IC50 values range from 7 to 159 μM after incubation for 72 h shown in Table 2.2. Etoposide was used as a standard drug and the complexes 2.1 and 2.4 showed good or similar activity to etoposide but the complexes do not extend the activity of the standard against the human cancer cells [35]. Ruthenium complexes sometimes display poor in vitro toxicity and good in vivo characteristics [36]. Some arene-ruthenium derivatives (2.6–3.0) of general formula [Ru( ɲ6p-cymene)(OCCH3NR)Cl] were synthesized in good yield and evaluated for their anticancer activity against human refractory metastatic prostate cancer (HRMPC) cell lines (Figure 2.8). From the data, it was verified that these complexes were successful in inhibiting the cell proliferation in both HRMPC PC-3 and DU-145 cells. The GI50 (μM) values for PC-3 and DU-145 cells using these complexes were less than 30 μM ranging from 4.25 to 19.68 μM and 5.48 to 20.93 μM, respectively [37]. Complex 2.7 displayed good result with values of 4.25 and 5.48 μM for PC-3 and DU-145. The GI50 values of cisplatin, [(ɲ6-p-cymene)RuII(L)Cl]Cl [38] and [Ru(ɲ6cymene)Cl(ɲ2-dppm)]PF6 [39], are shown in Table 2.3.

N N

N S

Ru N

N N

NHR

Ru

•2PF 6

NH

N O O

NHR •2PF 6

NH N

N

H

R = H(2.1); R = C2H5 (2.2)

S

N

H

O O

R = H(2.3); R = CH3 (2.4); R = C2H5 (2.5)

Figure 2.7  Structures of the complexes displaying poor in vitro toxicity and good in vivo characteristics.

369 ± 4.9

198 ± 4

101 ± 6

17.8 ± 2.5

65.9 ± 2.5

27.2 ± 1.2

2.2

2.3

2.4

2.5

Etoposide

24 h

2.1

IC50

Comp.

HCT-116

8.6 ± 2.1 18.3 ± 1.4

22.9 ± 1.5

8.6 ± 2.1

55.6 ± 3.6

159 ± 4

27.5 ± 3.3

72 h

13.4 ± 3.1

13.4 ± 3.1

67.4 ± 4.1

174 ± 6

324 ± 3.3

48 h

23.1 ± 1.2

19.5 ± 2.8

19.5 ± 2.8

97.0 ± 6.4

148 ± 3

39.2 ± 4.0

24 h

Caco-2

17.9 ± 1.3

15.2 ± 3.7

15.2 ± 3.7

76.4 ± 4.0

138 ± 3

31.0 ± 4.4

48 h

16.5 ± 2.0

6.6 ± 2.4

6.6 ± 2.4

58.9 ± 4.1

130 ± 3

22.2 ± 4.7

72 h

50.1 ± 1.5

212 ± 6

212 ± 6

221 ± 6

nd

220 ± 6

24 h

CCD-18Co

45.2 ± 1.1

136 ± 3

136 ± 3

179 ± 5

321 ± 5

153 ± 4

48 h

42.2 ± 1.0

114 ± 2

114 ± 2

167 ± 4

304 ± 4

145 ± 3

72 h

Table 2.2  IC50 values (μM) showing the anti-proliferative activity of all the complexes against the three human cells: 2 cancerous (HCT-116 and Caco-2) and one non-cancerous (CCD-18Co).

48  Advances in Metallodrugs

Chemotherapeutic Potential of Ruthenium  49

Ru

Cl O

N

R

Figure 2.8  Structures of the Ruthenium complexes.

Complex

2.6

2.7

2.8

2.9

3.0

OMe

OMe

R

Table 2.3  In vitro cytotoxicity of complexes 2.6–3.0 and related compounds against human refractory metastatic prostate cancer cell lines PC-3 and DU-145. SRB assay (GI50, μM) Compound

PC-3 cells (N = 3)

DU-145 cells (N = 3)

2.6

14.54 ± 1.30

21.64 ± 2.46

2.7

4.25 ± 0.42

5.48 ± 0.35

2.8

5.34 ± 0.17

5.65 ± 0.14

2.9

19.68 ± 0.65

20.93 ± 2.69

3.0

12.39 ± 1.51

14.31 ± 0.90

Cisplatin

>30

>30

[( ɲ6-p-cymene)RuII(L1)Cl]Cla

44.9

[( ɲ6-p-cymene)RuII(L2)Cl]Clb

>80

[Ru(ɲ 6-cymene) Cl( ɲ2-dppm)]PF6

1.9

[Ru(ɲ 6-cymene) Cl( ɲ2-dppmO)]PF6

>100

L1 = bis(3,5-dimethylyrazolyl)parabenzoic acid. L2 = bis(3,5-dimethylyrazolyl)metabenzoic acid.

a

b

50  Advances in Metallodrugs Another series of Ruthenium(II) complexes (3.1–3.8) showed potent cytotoxic effects against three cancer cell lines MCF-7 (breast), MDA-MB-468 (breast), and A431 (skin) (Figure 2.9). Cisplatin was taken as standard and the IC50 values of these complexes fall in the range of 4.5–31.71 μM. The complex 3.3 showed higher cytotoxic activity than the other complexes. The IC50 values of complex 3.3 against MCF-7, MDA-MB, and A431 are 4.5, 7.24 and 19.18 μM, respectively [40]. Novel Schiff base ligand-based Ruthenium(II) complexes (3.9–4.1) displayed interesting cytotoxic effects against HeLa and HEp2 tumor cells. These complexes were synthesized following Scheme 2.1. The IC50 values of the ligand against HeLa and Hep2 cell lines are 175 and 389 μM, respectively. These values revealed higher activity against HeLa cell line and lower against Hep2. The IC50 values of complexes against HeLa and Hep2 cell lines are 149 and 244 μM for complex 3.5; 123 and 212 μM for complex 3.6; and 87 and 149 μM for complex 3.7, respectively. With the coordination of the ruthenium metal the IC50 value of the ligand got decreased. The complex 4.1 showed better activity as compared with the ligand and other complexes and the results are shown in Table 2.4 [41]. Ruthenium(III) complexes have a great tendency for ligand exchange reactions with S-methylisothiosemicarbazone ligands and thus interact more quickly with the target molecules [42, 43]. A series of ruthenium(III) S-methylisothiosemicarbazone complexes (4.1–4.8) were synthesized and the structure of these complexes were analyzed by various analytical and spectral methods (Figure 2.10). The anticancer activity of the complexes (4.2–4.5) was checked against the human breast cancer cell line (MCF-7). The complexes 4.6 and 4.7 exhibited more activity than the standard drug cisplatin (Table 2.5). The complex 4.5 showed activity close to cisplatin and the complex 4.8 showed poor activity as compared to all the complexes [44]. EPh3 O N

Cl Ru Y Co N C HN R

E = P or S; Y = S or O; R = H, CH3 or C6H5

Figure 2.9  Structures of the ruthenium complexes supported by tridentate ligands derived from phenanthrenequinone and derivatives of thiosemicarbazide/carbazide.

Chemotherapeutic Potential of Ruthenium  51

N

HO N

S

+

[RuHCl(CO)(B)(EPh3)2]

H Methanol Reflux 4h

B Cl

OC Ru

O

N N S

H

Where B = PPh3/AsPh3/Py; E = PPh3/AsPh3 PPh3 (Triphenyl phosphne); AsPh3 (Triphenyl arsine); Py (Pyridine)

Scheme 2.1  Formation of the ruthenium(II) complexes synthesized from the Schiff base ligand 3-(benzothiazole-2-yliminomethyl)-napthalen-2-ol.

Table 2.4  The IC50 (μM) values for ruthenium(II) complexes (3.9–4.1) against selected cell lines. Compound

HeLa

HEp2

3.9

149

244

4.0

123

212

4.1

87

149

A three hexa-coordinated ruthenium(II) Schiff base complex of the type [RuCl(CO)(B)L] was synthesized (Figure 2.11). The ligand and the complex were tested for antitumor activity against the human cervical carcinoma cell line HeLa. The IC50 values (μM) of the ligand and complex was calculated using the MTT assay. The IC50 values calculated for the ligand and the complex were 52.3 and 31.6 μM, respectively. The complex exhibited higher activity than the ligand and the IC50 values calculated for the ligand and the complex are higher than the standard drug cisplatin with IC50 value of 16.7 μM [45].

52  Advances in Metallodrugs EPh3

R

O N

R O

Ru Cl

N N

H3CS E = P or As R = H, 5-Cl, 3-CH3O, C4H4 (Phenyl ring)

Figure 2.10  Structures of ruthenium(III) complexes (3.8–4.5) from the reactions between [RuCl3(EPh3)3] and bis(salicylaldehyde)-S-methylisothiosemicarbazone (H2L1-3)/bis (2-hydroxy-napthaldehyde)-S-methylisothiosemicarbazone (H2L4) ligands.

Table 2.5 IC50 values (μM) of ruthenium(III) complexes and cisplatin against the human breast cancer cell line (MCF-7). Complex

IC50 (μM)a

4.5

12.86 ± 0.3

4.6

7.24 ± 0.08

4.7

0.90 ± 0.1

4.8

21.19 ± 0.3

Cisplatin

12.33 ± 0.8

Fifty percent inhibitory concentration after exposure for 48 h in the MTT assay.

a

CO N

N

B

O Ru N HC

Cl

O

B = PPh3 or AsPh3 or Py

Figure 2.11  Structure of the ruthenium(III) complex synthesized from the Schiff base ligand and [RuHCl(CO)(EPh3)2(B)] where E = P or As or Py.

Chemotherapeutic Potential of Ruthenium  53 α[Ru(azpy)2Cl2] is among the most cytotoxic Ru(II) compound reported in the literature [46–48]. α[Ru(azpy)2Cl2] and doxorubicin show the strong activity against all the cancer cell lines and having the lowest IC50 values compared to other compounds. Six complexes of Ru(II)-bis(arylimino)pyridine complexes, synthesized from [RuCl3(L1)](H2O)] and several bidentate co-ligands that include1,10-phenanthroline(phen), 2,2’dipyridyl (bpy), 2-(phenylazo)pyridine (azpy), 2-(phenylazo)-3-methylpyridine (3mazpy), 2-(tolylazo)pyridine (tazpy), and 2-picolinate (pic) (Figure 2.12) have been reported as anticancer agents. The in vitro cytotoxic effects of these new Ru(II) complexes (4.9–5.8) were checked in comparison with the parent Ru(III) complex. The anticancer activity of these complexes was tested against seven human cell lines, namely, A498, EVSA-T, H226, IGROV, M19, MCF-7, and WIDR. The reference compounds used were cisplatin and doxorubicin. The IC50 values were checked after the continuous exposure of the cells to the compounds for 120 h (Table 2.6). These compounds show moderate to high cytotoxic effects and some compounds showed better activity than the cisplatin. The IC50 values show that the complexes of the type Ru(II)L1(LL) have lower values (higher cytotoxic activity) (in the range of 0.4–6 μM) than the parent [RuIIICl3(L1)](H2O) and [RuIII(tpy) (azpy)Cl.5H2O [49]. A series of RuII arene complexes were synthesized from the mono and bidentate N-donor ligands (5.9–6.3) (Scheme 2.2). The ligands and the

L

+

Ru N

N

N

N

L

N N

N

L L

Cl

N N

bpy

N

Phen azpy

L bidentate ligand

N

N N

N

N N

N -O

3mazpy

tazpy

O pic

Figure 2.12  Structures of Ru(II) derivatives synthesized from [RuCl3(L1)](H2O) (L1: 2,6-bis(2,4,6-trimethylphenyliminomethyl)pyridine)with several bidentate co-ligands.

15.1

79.6

39.3

18.0

19.2

9.7

5.7

4.1

2.9

0.3

7.5

0.10

4.9b

5.0

5.1c

5.2

5.3

5.4

5.5

5.6

5.7

5.8d

Cisplatin

DOX

0.015

1.4

0.06

0.4

0.5

0.6

0.4

0.8

1.4

11.4

67.0

11.2

EVSA-T

0.37

10.9

0.5

1.8

3.0

3.2

3.6

9.1

13.4

33.6

63.7

15.2

H226

0.11

0.6

0.3

2.0

6.5

2.8

1.9

3.7

26.7

64.8

90.1

12.2

IGROV

0.03

1.9

0.06

0.4

0.6

0.9

1.0

1.5

2.0

14.6

68.5

12.2

M19

0.02

2.3

0.3

0.8

1.7

1.5

1.0

2.2

4.2

30.5

64.0

17.1

HCF-7

0.02

3.2

0.3

1.4

2.2

2.3

1.5

3.1

10.5

51.0

79.0

14.5

WIDR

a

A498 human renal carcinoma cell line; EVSA-T human breast cancer cell line; H226 human non-small cell lung carcinoma cell line; IGROV human ovarian carcinoma cell line; M19 human melanoma carcinoma cell line; HCF-17 human breast adenocarcinoma cell line; WIDR human colon adenocarcinoma cell line. bRef. 50, cRef. 51, dRef. 48.

A498

Compounds

Cell linea, IC50 (μM)

Table 2.6  In vitro cytotoxic effect of the synthesized compounds incubated for 120 h; the uncertainties are estimated to be 0.01–0.03 μM.

54  Advances in Metallodrugs

Chemotherapeutic Potential of Ruthenium  55

N

H2N

N

N H

5.9

H N

H N

O 6.2

OH

N H 6.4

H

O

6.3

H

6.0 or 6.1 MeOH RT

5.9 MeOH RT

O Ru

N H 6.1

6.0

N

Cl

OH

O

H

Cl

O

O

O

Cl

Ru Cl 6.5 O

N R

O

6.6

O

OH

R D

Cl

Cl

NH

Cl

PF6

E NH4PF6 MeOH reflux O

Ru N

H H N

6.7 Cl

NH

Ru

L

Cl

O

L

H N

Cl

Ru

L

NH4PF6 MeOH reflux

Ru

O

PF6

H

O2 -H2O

N Cl

N 6.8

PF6

Ru N 6.9

Scheme 2.2  Molecular structures of ligands 5.9–6.3 and synthesized complexes 6.4–6.9.

synthesized complexes 6.4–6.9 were tested for cytotoxicity against various human tumor cell lines, i.e., 8505C, MCF-7, SW-480, and 518A2. The cytotoxic effect was checked after exposure of the cells to the compounds for 90 h using SRB assay. Among the ligands tested, 6.0 and 6.3 showed cytotoxic effect compared to the Ru(II) complexes. The IC50 values obtained for the complexes were higher than the reference compound cisplatin [52]. The complex 6.5 was more effective than 6.6, while complexes 6.4, 6.7, and

56  Advances in Metallodrugs 6.9 revealed no anticancer activity (Table 2.7). The anticancer activity of complex 6.5 is associated to inhibition of cell proliferation and subsequent caspase-dependent apoptosis [53]. Half sandwich Ru(II) complexes (7.5–7.9) were prepared from the azo containing Schiff base ligands (7.0–7.4) (Figure 2.13) which were in turn synthesized from (E)-((4-ethylphenyl)-diazenyl)-2-hydroxybenzaldehyde Table 2.7  IC50 values (μM) of the ligands (5.9–6.3) and complexes (6.4–6.9) against various human cancer cell lines. Compounds

8505C

MCF-7

SW-480

518A2

5.9

>100

>100

>100

>100

6.0

79.6 ± 4.5

20.2 ± 3.5

91.1 ± 4.9

>100

6.1

>100

55.8 ± 0.3

>100

>100

6.2

>100

>100

>100

>100

6.3

58.2 ± 11.6

>100

44.1 ± 5.1

40.5 ± 4.5

6.4

>100

>100

>100

>100

6.5

69.1 ± 2.5

36.3 ± 2.3

94.1 ± 18.3

97.7 ± 3.2

6.6

90.2 ± 13.9

42.5 ± 0

>100

>100

6.7

>100

>100

>100

>100

6.9

>100

>100

>100

>100

Cisplatin

4.8 ± 0.1

2.2 ± 0.2

3 ± 0.4

2 ± 0.4

Cl OH N

N

O N

N

5.1, R = p-CH3 5.2, R = p-OCH3 5.3, R = O-OCH3 5.4, R = 3,4-OCH3 5.5, R = 2,4,6-CH3

R

N

Ru N

5.6, R = p-CH3 5.7, R = p-OCH3 5.8, R = O-OCH3 5.9, R = 3,4-OCH3 6.0, R = 2,4,6-CH3

R

Figure 2.13  Structures of the Ru(II) complexes (5.6–6.0) synthesized from the azo containing Schiff base ligands (5.1–5.5).

Chemotherapeutic Potential of Ruthenium  57 and aniline derivatives. The antiproliferative activity of these synthesized compounds was checked against H2126, PC3, and MCF-7 human cancer cell lines. Both acute and chronic effects were checked at 200 μM concentration. The IC50 values (μM) calculated for these compounds are shown in Table 2.8. The IC50 values revealed that some of the compounds inhibited cell proliferation in the acute phase (first 4 h) and did not reveal any effect in the chronic phase (first 48 h). Some compounds show more activity against the prostate cancer cell lines but did show any effect against the lung and breast cancer cell lines. Out of the 10 compounds tested, 7.2 and 7.3 were most effective. The phenyl ring in the compounds have methyl or methoxy groups with reference to the azomethine bond and as the number of these groups increase, anti-cancer activity moderately decreased, which may be due to the steric effect of these groups [54]. Some new cyclometallated Ru(II) complexes of 3-acetyl-7-methoxycoumarin-4N-substituted thiosemicarbazones were synthesized (Figure 2.14). The anticancer potential of the metal precursor [RuHClCO(PPh3)3] and synthesized compounds was checked against MCF-7 (human breast cancer) and A549 (human lung cancer) cell lines by using MTT assay. The IC50 values (μM) of these complexes revealed that they are more effective than the other reported Ru(II) complexes [53, 55–60]. The non-toxic nature of these compounds was checked against non-cancerous cell line HaCaT (human normal keratinocyte). Cisplatin was used as a positive control. The IC50 values of these compounds revealed that they were cytotoxic against these cells. The IC50 values of coumarin attached thiosemicarbazones (8.0– 8.3) and Ru(II) complexes (8.4–8.7) against these cancer cell lines were lower than that of cisplatin, signifying their good activity over cisplatin (Table 2.9). The cytotoxic effect of Ru(II) complexes was higher than that of cisplatin and their parent ligands. The coordination of ligands to the Ru(II) metal ion considerably increased the cytotoxic effect of the complexes six times over ligands and eight times over cisplatin against both the cell lines. Among all the four complexes Ru(II), complex 8.6 showed higher activity against both the cell lines because of having the more electron donating ethyl group at N-terminal nitrogen followed by complex 8.5 (NH-Me), complex 8.4 (NH-H), and then complex 8.7 having electron withdrawing phenyl group at terminal nitrogen atom [61]. Anticancer activity of a new series of ketone −N4 substituted thiosemicarbazone (TSC) compounds (8.9–9.6) and their corresponding [(ƞ6-p-­ cymene)RuII(TSC)Cl]+/0 complexes (9.7–10.5) (Scheme 2.3) was evaluated against SGC-7901 (human gastric cancer), BEL-7404 (human liver cancer), and HEK-293T (noncancerous) cell lines. Cisplatin, oxaliplatin, and carboplatin were taken as reference compounds. The IC50 values of most

4 h - acute

6.008

6.062

5.429

5.277

5.307

6.132

6.246

5.682

4.8

5.47

5.197

Compound

7.0

7.1

7.2

7.3

7.4

7.5

7.6

7.7

7.8

7.9

Positive control (5-FU)

1.215

10.51

9.242

9.806

10.81

9.39

9.413

9.305

9.353

9.288

8.695

48 h - chronic

MCF7 (breast cancer)

IC50 (μM)

3.637

3.843

2.868

3.381

3.648

3.481

3.122

2.85

2.763

3.456

3.467

4 h - acute

1.204

8.975

8.017

7.362

7.631

7.517

8.235

8.944

6.118

7.114

8.026

48 h - chronic

PC3 (prostate cancer)

2.872

3.211

3.121

3.17

3.425

3.177

3.18

2.9

3.201

3.344

3.063

4 h - acute

2.538

4.338

4.537

4.994

4.219

4.412

4.372

4.714

5.021

5.115

4.94

48 h - chronic

H2126 (lung cancer)

Table 2.8  Antiproliferative activity of the ligands (5.1–5.5), Ru(II) complexes (5.6–6.0), and positive control (5-FU).

58  Advances in Metallodrugs

Chemotherapeutic Potential of Ruthenium  59 O

O R

N

O

N P O Ru

O

O P R=H R=CH3 R=C2H5 R=C6H5

OC

R=H R=CH3 R=C2H5 R=C6H5

8.0 8.1 8.2 8.3

R

8.4 8.5 8.6 8.7

Figure 2.14  Structures of the synthesized ligands 8.0–8.3 and their ruthenium(II) complexes (8.4–8.7).

Table 2.9  The IC50 values of the ligands (8.0–8.3), [RuHClCO(PPh3)3], and new Ru(II) complexes (8.4–8.7) against MCF-7 (human breast cancer cell line), A549 (human lung cancer cell line), and HaCaT (non-cancerous keratinocyte cells) for 48 h. IC50 values (μM) Compounds

MCF-7

A549

HaCaT

Cisplatin

16.79 ± 0.08

15.10 ± 0.05

>40

8.0

13.06 ± 0.29

12.64 ± 0.17

>40

8.1

12.12 ± 0.32

12.12 ± 0.16

>40

8.2

11.27 ± 0.21

11.63 ± 0.15

>40

8.3

13.11 ± 0.25

13.83 ± 0.18

>40

[RuHClCO(PPh3)3]

20.10 ± 0.18

15.96 ± 0.21

>40

8.4

2.86 ± 0.17

2.96 ± 0.07

>40

8.5

2.62 ± 0.07

2.93 ± 0.07

>40

8.6

2.53 ± 0.10

2.37 ± 0.04

>40

8.7

3.02 ± 0.05

3.05 ± 0.12

>40

of the TSC ligands were relatively higher and are considered as inactive against both the cancer cell lines. Among the TSC ligands, 9.0 showed the highest cytotoxic effect with an IC50 value of 39 μM against SGC7901 cancer cell lines, 9.3 displayed the highest activity with an IC50 value around 37 μM against the BEL-7404 cancer cell lines. The Ru-arene complexes displayed increased anticancer activity with an IC50 value around

60  Advances in Metallodrugs R1

N R2

S N H

N H

R3

TSC Complex R1 R2 R3

+

[(n6-p-cymene)RuCl2]2

TCSs (8.8-9.6)

L Cl

Ru S N 9.7-10.5

+/0

8.8

9.7

Me Me

8.9

9.8

Me Me Me

9.0

9.9

Me Me Ph

9.1

10.0

Ph Me

9.2

10.1

Ph Me Me

9.3

10.2

Ph Me Ph

9.4

10.3

Ph Ph

9.5

10.4

Ph Ph Me

9.6

10.5

Ph Ph Ph

H

H

H

Scheme 2.3 Synthetic route of the TSC compounds (8.8–9.6) and their corresponding [(ƞ6-p-cymene)RuII(TSC)Cl]+/0 complexes (9.7–10.5).

(16–48 μM) as compared to that of free ligands against both the cancer cell lines (Table 2.10). The complexes showed comparable activity to cisplatin and oxaliplatin (16–32 μM) and showed more cytotoxic effect than carboplatin against BEL-7404 cancer cell line signifying the good cytotoxicity of these complexes. The compounds (9.9, 10.2, and 10.5) with the N4 substitution of the phenyl ring displayed the higher anticancer activity against both the cancer cell lines as compared to that of H and methyl substituted compounds. Cytotoxicity of the compounds was also checked against HEK-293T (non-cancerous) cell lines, and in most of the compounds, the cytotoxicity was comparable for both the cancerous and the normal cell lines [62]. New ruthenium half sandwiched complexes (10.6–11.0) containing (N,N)-bound picolinamide and quinaldamide ligands were synthesized (Figure 2.15) and evaluated for their cytotoxicity against HT-29 (human colon adenocarcinoma), MCF-7 (human breast adenocarcinoma) cell lines over a 5-day exposure and a further 1 h for MCF-7 cells. The cytotoxicities of the ruthenium picolinamide complexes (10.6–10.9) were in the order of 10.9>10.8~10.7 >10.6 for both the cell lines (Table 2.11). The quinaldamide complex (11.0) and picolinamide 10.7 complex showed similar activity. The complex 10.9 was the most cytotoxic compared to other complexes. All the complexes showed low activity towards MCF-7 cells after a 1-h exposure as compared to 5-day exposure. The complex 10.9 displayed

SGC-7901

>100

>100

38.6 ± 4.3

62.2 ± 6.1

>100

>100

>100

>100

>100

TSC

8.8

8.9

9.0

9.1

9.2

9.3

9.4

9.5

9.6

IC50 (μM)

45.4 ± 4.1

>100

50.1 ± 4.9

37.1 ± 0.1

>100

>100

>100

96.1 ± 9.7

>100

BEL-7404

>100

>100

51 ± 6.3

>100

68 ± 5.9

41 ± 4.1

97 ± 8.3

>100

>100

HEK-293T

17.0 ± 2.9 6.7± 0.4 11 ± 0.8 39 ± 2.0

Cisplatin Oxaliplatin Carboplatin

34.4 ± 3.1

47.7 ± 1.3

46.7 ± 0.6

43.4 ± 9.3

28.6 ± 3.5

17.5 ± 1.6

39.9 ± 2.6

30.8 ± 5.7

SGC-7901

10.5

10.4

10.3

10.2

10.1

10.0

9.9

9.8

9.7

Complex

IC50 (μM)

70 ± 9.5

20 ± 0.6

23.1 ± 2.6

17.1 ± 4.6

21.8 ± 4.4

24.5 ± 2.2

15.9 ± 2.2

29.7 ± 0.7

29.1 ± 7.1

18.2 ± 3.2

28.2 ± 0.6

32.0 ± 3.1

BEL-7404

44 ± 3.7

2 ± 0.2

10 ± 0.7

10 ± 2.0

17 ± 2.2

20 ± 1.3

17 ± 4.3

33 ± 0.4

46 ± 2.5

25 ± 0.9

22.4 ± 3.5

>100

HEK-293T

Table 2.10  IC50 values (μM) of TSC compounds (8.8–9.6), complexes (9.7–10.5), cisplatin, oxaliplatin, and carboplatin against SGC-7901, BEL-7404, and HEK-293T cell lines.

Chemotherapeutic Potential of Ruthenium  61

62  Advances in Metallodrugs

Ru

N

Cl

N

N

N R

O

Ru

Cl Cl

O

10.6 R = 2'-Cl 10.7 R = 3'-Cl 10.8 R = 2',4'-Cl 10.9 R = 2',5'-Cl

Cl 11.0

Figure 2.15  Structures of Ruthenium-para-cymene Picolinamide complexes (10.6–10.9) and Ruthenium-para-cymene-Quinaldamide complex (11.0).

Table 2.11  IC50 values for complexes (10.6–11.0) and cisplatin against HT-29 and MCF-7 cancer cell lines. Compound

HT-29 IC50/μMa

MCF-7 IC50/μMa

MCF-7 IC50/μMb

10.6

33 ± 7

35 ± 14

184 ± 2

10.7

13 ± 3

11.2 ± 0.7

–

10.8

16 ± 3

11.5 ± 0.9

64 ± 17

10.9

5.9 ± 0.8

5±1

32 ± 15

11.0

11.5 ± 0.7

13 ± 3

–

Cisplatin

10 ± 3

3±1

53± 8

incubated for 5 days; bincubated for 1 h.

a

cytotoxicity after a 1-h exposure, and after the 5-day exposure of the same cell line, this complex showed the higher cytotoxic effect compared to cisplatin. This showed that complex 10.9 is more potent drug than cisplatin. The complex 10.9 was further tested on MCF-7 cells in a hypoxic environment and this complex retained its activity against hypoxic cells. This complex has the potential to eliminate both the aerobic and hypoxic fraction of cell cells and has the capability to be used for further studies [63]. A new series of binuclear ruthenium(II) complexes (11.1–11.4) were synthesized (Figure 2.16) and tested in vitro against HeLa (cervical carcinoma), SW620 (metastatic colorectal adenocarcinoma), A549 (lung ­adeno-carcinoma), MCF-7 (breast adenocarcinoma), and WI-38 (human lung fibroblast) cell lines. All the complexes showed good anticancer

Chemotherapeutic Potential of Ruthenium  63 X

HC N N

Cl O Ru

NH2

R

Cl Ru

O H2O

11.1 Cl

N

11.2 H

N

11.3 Br

CH

11.4 NO2

X

Figure 2.16  Structures of Ruthenium(II) complexes with Schiff bases derived from 5-chlorosalicyladehyde and 2-aminopyridine and its 5-substituted salicylideneimine homologues.

Table 2.12  IC50 values (μM) for ruthenium(II) complexes (11.1–11.4) against HeLa (cervical carcinoma), SW620 (metastatic colorectal adenocarcinoma), A549 (lung adenocarcinoma), and MCF-7 (breast adenocarcinoma) cell lines. IC50 (μM) Compound SW620

A549

MCF-7

HeLa

W138

11.1

3.26 ± 0.17 1.5 ± 0.79

4.59 ± 0.42 2.23 ± 0.82 2.74 ± 0.48

11.2

1.99 ± 0.56 0.68 ± 0.88

4.09 ± 0.78 1.66 ± 0.48 2.51 ± 98.88

11.3

3.00 ± 0.21 1.38 ± 0.41

4.70 ± 0.16 2.26 ± 1.09 4.01 ± 2.08

11.4

5.24 ± 0.86 26.85 ± 24.9 8.13 ± 0.95 5.80 ± 4.31 31.62 ± 69.51

activity against these cancer cell lines with low IC50 values (Table 2.12). The compound 11.2 exhibited strong activity against A549 with extremely low IC50 = 0.68 μM as compared to other compounds. The anticancer activity of these compounds is in the order of 11.2>11.1~11.3>11.4 revealing an effect of substituent on aldehyde ring of Schiff base [64].

2.3 Conclusion Since cancer is a complex disease, different treatment strategies are required to curb or treat any particular cancer. Some cancers require only chemotherapy while others require combination therapy such as surgery

64  Advances in Metallodrugs with chemotherapy or radiotherapy. Although tremendous advances have been made based on latest technologies, there is no ideal treatment for cancer. Therefore, efforts are to be made in every direction to bring new chemotherapeutic agents or novel treatment strategies to augment the currently used medication for cancer. A plethora of chemotherapeutic agents are already available in the market but there are growing concerns about their limited tumor specificity and off-target toxicity. Drug resistance is another big concern. The success of Platinum-based chemotherapeutic agents paved the way for development of other similar metal-based agents in which Ruthenium-based complexes stood out to be the best alternatives with enhanced activity and limited toxicity. Although Ruthenium metal finds nowhere any mention of its biological role in microbiology texts, complexation of this metal with diverse ligands exposed its striking biological applications. Undoubtedly, Schiff base ruthenium complexes have emerged as promising molecules against cancer. It is because of the fact that the ligand-exchange reactions of ruthenium complexes are slow, resulting in the delayed defragmentation of these compounds before they reach to their biological targets. Besides that, selective accumulation in tumor cells, unique DNA binding modes and activation by reduction in more hypoxic and acidic tumor cells are characteristic of ruthenium complexes. Therefore, there is a great hope that Ruthenium-based chemotherapeutic agents could be developed in the near future.

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66  Advances in Metallodrugs from size exclusion chromatography-ICP-MS and ESI-MS. J. Anal. At. Spectrom., 25, 305, 2010. 20. Antonarakis, E.S. and Emadi, A., Ruthenium-based chemotherapeutics: Are they ready for prime time? Cancer Chemother. Pharmacol., 66, 1–9, 2010. 21. Frei, A., Rubbiani, R., Tubafard, S., Blacque, O., Anstaett, P., Felgenträger, A., Maisch, T., Spiccia, L., Gasser, G., Synthesis, characterization, and biological evaluation of new RU(ii) polypyridyl photosensitizers for photodynamic therapy. J. Med. Chem., 57, 7280–7292, 2014. 22. Bergamo, A. and Sava, G., Ruthenium anticancer compounds: Myths and realities of the emerging metal-based drugs. Dalton Trans., 40, 7817, 2011. 23. Baile, M.B., Kolhe, N.S., Deotarse, P.P., Jain, A.S., Kulkarni, A.A., Metal Ion Complex -Potential Anticancer Drug- A Review. Int. J. Pharm. Res. Rev. IJPRR, 4, 59–66, 2015. 24. Florea, A.M. and Büsselberg, D., Cisplatin as an Anti-Tumor Drug: Cellular Mechanisms of Activity, Drug Resistance and Induced Side Effects. Cancers (Basel), 3, 1351–1371, 2011. 25. Barry, N.P.E. and Sadler, P.J., Exploration of the medical periodic table: Towards new targets. Chem. Commun., 49, 5106, 2013. 26. Ndagi, U., Mhlongo, N., Soliman, M.E., Metal complexes in cancer therapy–an update from drug design perspective. Drug Des. Devel. Ther., 11, 599–616, 2017. 27. Rajapakse, C.S.K., Martínez, A., Naoulou, B., Jarzecki, A.A., Suárez, L., Deregnaucourt, C., Sinou, V., Schrével, J., Musi, E., Ambrosini, G., Schwartz, G.K., Sánchez-Delgado, R.A., Synthesis, characterization, and in vitro antimalarial and antitumor activity of new ruthenium(II) complexes of chloroquine. Inorg. Chem., 48, 1122–1131, 2009. 28. Lallan, M. and Kumari, N., Approaching Cancer Therapy with Ruthenium Complexes by Their Interaction with DNA, in: Ruthenium Chemistry, pp. 259–303, Pan Stanford, New York, 2018. 29. Sinha, D., Tiwari, A.K., Singh, S., Shukla, G., Mishra, P., Chandra, H., Mishra, A.K., Synthesis, characterization and biological activity of Schiff base analogues of indole-3-carboxaldehyde. Eur. J. Med. Chem., 43, 160–165, 2008. 30. Guo, Z. and Sadler, P.J., Medicinal Inorganic Chemistry: Introduction. Chem. Rev., 99, 2201–2204, 1999. 31. Kalaivani, P., Prabhakaran, R., Poornima, P., Dallemer, F., Vijayalakshmi, K., Padma, V.V., Natarajan, K., Versatile coordination behavior of salicylaldehydethiosemicarbazone in ruthenium(II) carbonyl complexes: Synthesis, spectral, X-ray, electrochemistry, DNA binding, cytotoxicity, and cellular uptake studies. Organometallics, 31, 8323–8332, 2012. 32. Jayanthi, E., Anusuya, M., Bhuvanesh, N.S.P., Khalil, K.A., Dharmaraj, N., Synthesis and characterization of ruthenium(II) hydrazone complexes as anticancer chemotherapeutic agents: In vitro DNA/BSA protein binding and cytotoxicity assay. J. Coord. Chem., 68, 3551–3565, 2015. 33. Gichumbi, J.M., Friedrich, H.B., Omondi, B., Singh, M., Naicker, K., Chenia, H.Y., Synthesis, characterization, and cytotoxic and antimicrobial activities

Chemotherapeutic Potential of Ruthenium  67 of ruthenium(II) arene complexes with N,N-bidentate ligands. J. Coord. Chem., 69, 3531–3544, 2016. 34. Ejidike, I.P. and Ajibade, P.A., Synthesis, characterization, and in vitro anti­ oxidant and anticancer studies of ruthenium(III) complexes of symmetric and asymmetric tetradentate Schiff bases. J. Coord. Chem., 68, 2552–2564, 2015. 35. Beckford, F.A., Thessing, J., Shaloski, M., Mbarushimana, P.C., Brock, A., Didion, J., Woods, J., Gonzalez-Sarrías, A., Seeram, N.P., Synthesis and characterization of mixed-ligand diimine-piperonal thiosemicarbazone complexes of ruthenium(II): Biophysical investigations and biological evaluation as anticancer and antibacterial agents. J. Mol. Struct., 992, 39–47, 2011. 36. Scolaro, C., Bergamo, A., Brescacin, L., Delfino, R., Cocchietto, M., Laurenczy, G., Geldbach, T.J., Sava, G., Dyson, P.J., In Vitro and in Vivo Evaluation of Ruthenium(II)–Arene PTA Complexes. J. Med. Chem., 48, 4161–4171, 2005. 37. Lin, T.H., Das, K., Datta, A., Leu, W.-J., Hsiao, H.-C., Lin, C.-H., Guh, J.-H., Huang, J.-H., Synthesis and characterization of ruthenium compounds incorporating keto-amine ligands. The applications of catalytic transfer hydrogenation and cancer cell inhibition. J. Organomet. Chem., 807, 22–28, 2016. 38. Khan, R.A., Arjmand, F., Tabassum, S., Monari, M., Marchetti, F., Pettinari, C., Organometallic ruthenium(II) scorpionate as topo IIα inhibitor; in vitro binding studies with DNA, HPLC analysis and its anticancer activity. J. Organomet. Chem., 771, 47–58, 2014. 39. Das, S., Sinha, S., Britto, R., Somasundaram, K., Samuelson, A.G., Cytotoxicity of half sandwich ruthenium(II) complexes with strong hydrogen bond acceptor ligands and their mechanism of action. J. Inorg. Biochem., 104, 93–104, 2010. 40. Anitha, P., Chitrapriya, N., Jang, Y.J., Viswanathamurthi, P., Synthesis, characterization, DNA interaction, antioxidant and anticancer activity of new ruthenium(II) complexes of thiosemicarbazone/semicarbazone bearing 9,10-phenanthrenequinone. J. Photochem. Photobiol. B Biol., 129, 17–26, 2013. 41. Sathiyaraj, S., Butcher, R.J., Jayabalakrishnan, C., Synthesis, characterization, DNA interaction and in vitro cytotoxicity activities of ruthenium(II) Schiff base complexes. J. Mol. Struct., 1030, 95–103, 2012. 42. Clarke, M.J., Bitler, S., Rennert, D., Buchbinder, M., Kelman, A.D., Reduction and Subsequent Binding of Ruthenium Ions Catalyzed by Subcellular Components. J. Inorg. Biochem., 12, 79–87, 1980. 43. Clarke, M.J., Ruthenium metallopharmaceuticals. Coord. Chem. Rev., 232, 69–93, 2002. 44. Prakash, G., Manikandan, R., Viswanathamurthi, P., Velmurugan, K., Nandhakumar, R., Ruthenium(III) S-methylisothiosemicarbazone Schiff base complexes bearing PPh3/AsPh3 coligand: Synthesis, structure and biological investigations, including antioxidant, DNA and protein interaction,

68  Advances in Metallodrugs and in vitro anticancer activities. J. Photochem. Photobiol. B Biol., 138, 63–74, 2014. 45. Raja, G., Butcher, R.J., Jayabalakrishnan, C., Studies on synthesis, characterization, DNA interaction and cytotoxicity of ruthenium(II) Schiff base complexes. Spectrochim. Acta Part A Mol. Biomol. Spectrosc., 94, 210–215, 2012. 46. Hotze, A.C.G., Caspers, S.E., de Vos, D., Kooijman, H., Spek, A.L., Flamigni, A., Bacac, M., Sava, G., Haasnoot, J.G., Reedijk, J., Structure-dependent in vitro cytotoxicity of the isomeric complexes [Ru(L)2Cl2] (L = o-tolylazopyridine and 4-methyl-2-phenylazopyridine) in comparison to [Ru(azpy)2Cl2]. JBIC J. Biol. Inorg. Chem., 9, 354–364, 2004. 47. Hotze, A.C.G., van der Geer, E.P.L., Kooijman, H., Spek, A.L., Haasnoot, J.G., Reedijk, J., Characterization by NMR Spectroscopy, X-ray Analysis and Cytotoxic Activity of the Ruthenium(II) Compounds. Eur. J. Inorg. Chem., 13, 2648–2657, 2005. 48. Hotze, A.C.G., Bacac, M., Velders, A.H., Jansen, B.A.J., Kooijman, H., Spek, A.L., Haasnoot, J.G., Reedijk, J., New ctytotoxic and water soluble Bis(2phenylazopyridine) ruthenium (II) complexes. J. Med. Chem., 46, 1743– 1750, 2003. 49. Garza-Ortiz, A., Uma Maheswari, P., Siegler, M., Spek, A.L., Reedijk, J., A  new family of Ru(ii) complexes with a tridentate pyridine Schiff-base ligand and bidentate co-ligands: Synthesis, characterization, structure and in vitro cytotoxicity studies. New J. Chem., 37, 3450, 2013. 50. Garza-Ortiz, A., Maheswari, P.U., Siegler, M., Spek, A.L., Reedijk, J., Ruthenium(III) Chloride Complex with a Tridentate Bis(arylimino)pyridine Ligand: Synthesis, Spectra, X-ray Structure, 9-Ethylguanine Binding Pattern, and In Vitro Cytotoxicity. Inorg. Chem., 47, 6964–6973, 2008. 51. Corral, E., Hotze, A.C.G., Tooke, D.M., Spek, A.L., Reedijk, J., Ruthenium polypyridyl complexes Containing the bischelating ligand 2,2′-azobispyridine. Synthesis, characterization and crystal structures. Inorganica Chim. Acta, 359, 830–838, 2006. 52. Song, H., Kaiser, J.T., Barton, J.K., Crystal structure of Δ-[Ru(bpy)2dppz]2+ bound to mismatched DNA reveals side-by-side metalloinsertion and intercalation. Nat. Chem., 4, 615–620, 2012. 53. Richter, S., Singh, S., Draca, D., Kate, A., Kumbhar, A., Kumbhar, A.S., Maksimovic-Ivanic, D., Mijatovic, S., Lönnecke, P., Hey-Hawkins, E., Antiproliferative activity of ruthenium(II) arene complexes with mono- and bidentate pyridine-based ligands. Dalton Trans., 45, 13114–13125, 2016. 54. İnan, A., İkiz, M., Tayhan, S.E., Bilgin, S., Genç, N., Sayın, K., Ceyhan, G., Köse, M., Dağ, A., İspir, E., Antiproliferative, antioxidant, computational and electrochemical studies of new azo-containing Schiff base ruthenium(II) complexes. New J. Chem., 42, 2952–2963, 2018. 55. Huang, H., Zhang, P., Chen, Y., Qiu, K., Jin, C., Ji, L., Chao, H., Synthesis, characterization and biological evaluation of labile intercalative ruthenium(II)

Chemotherapeutic Potential of Ruthenium  69 complexes for anticancer drug screening. Dalton Trans., 45, 13135–13145, 2016. 56. Chow, M.J., Licona, C., Yuan Qiang Wong, D., Pastorin, G., Gaiddon, C., Ang, W.H., Discovery and Investigation of Anticancer Ruthenium–Arene SchiffBase Complexes via Water-Promoted Combinatorial Three-Component Assembly. J. Med. Chem., 57, 6043–6059, 2014. 57. Kalaivani, P., Prabhakaran, R., Poornima, P., Dallemer, F., Vijayalakshmi, K., Padma, V.V., Natarajan, K., Versatile Coordination Behavior of Salicylaldehydethiosemicarbazone in Ruthenium(II) Carbonyl Complexes: Synthesis, Spectral, X-ray, Electrochemistry, DNA Binding, Cytotoxicity, and Cellular Uptake Studies. Organometallics, 31, 8323–8332, 2012. 58. Jayanthi, E., Kalaiselvi, S., Padma, V.V., Bhuvanesh, N.S.P., Dharmaraj, N., Solvent assisted formation of ruthenium(III) and ruthenium(II) hydrazone complexes in one-pot with potential in vitro cytotoxicity and enhanced LDH, NO and ROS release. Dalton Trans., 45, 1693–1707, 2016. 59. Jeyalakshmi, K., Haribabu, J., Bhuvanesh, N.S.P., Karvembu, R., Halfsandwich RuCl 2 (η 6 -p-cymene) core complexes containing sulfur donor aroylthiourea ligands: DNA and protein binding, DNA cleavage and cytotoxic studies. Dalton Trans., 45, 12518–12531, 2016. 60. Chen, L., Peng, F., Li, G., Jie, X., Cai, K., Cai, C., Zhong, Y., Zeng, H., Li, W., Zhang, Z., Chen, J., The studies on the cytotoxicity in vitro, cellular uptake, cell cycle arrest and apoptosis-inducing properties of ruthenium methylimidazole complex [Ru(MeIm)4(p-cpip)]2+. J. Inorg. Biochem., 156, 64–74, 2016. 61. Kalaiarasi, G., Jeya Rajkumar, S.R., Dharani, S., Małecki, J.G., Prabhakaran, R., An investigation on 3-acetyl-7-methoxy-coumarin Schiff bases and their Ru(II) metallates with potent antiproliferative activity and enhanced LDH and NO release. RSC Adv., 8, 1539–1561, 2018. 62. Su, W., Qian, Q., Li, P., Lei, X., Xiao, Q., Huang, S., Huang, C., Cui, J., Synthesis, Characterization, and Anticancer Activity of a Series of Ketone-N 4-Substituted Thiosemicarbazones and Their Ruthenium(II) Arene Com­ plexes. Inorg. Chem., 52, 12440–12449, 2013. 63. Almodares, Z., Lucas, S.J., Crossley, B.D., Basri, A.M., Pask, C.M., Hebden, A.J., Phillips, R.M., McGowan, P.C., Rhodium, Iridium, and Ruthenium Half-Sandwich Picolinamide Complexes as Anticancer Agents. Inorg. Chem., 53, 727–736, 2014. 64. Kahrović, E., Zahirović, A., Kraljević Pavelić, S., Turkušić, E., Harej, A., In vitro anticancer activity of binuclear Ru(II) complexes with Schiff bases derived from 5-substituted salicylaldehyde and 2-aminopyridine with notably low IC50 values. J. Coord. Chem., 70, 1683–1697, 2017.

3 Role of Metallodrugs in Medicinal Inorganic Chemistry Manish Kumar, Gyanendra Kumar, Arun Kant and Dhanraj T. Masram* Department of Chemistry, University of Delhi, Delhi, India

Abstract

Over the past decades, metals and metal-based drugs were significantly used in the treatment of several diseases but it cannot be a clear distinction between the toxic doses and the therapeutic which is a major challenge. Metal ions are required for many critical functions in humans. The medicinal uses of metal-based drugs are of increasing clinical and commercial importance. With the discovery of cisplatin in 1960 by Barnett Rosenberg, the study in these areas has been expanded exponentially but the use of transition metal-based compounds other than platinum has also attracted attention. However, the efficacy of platinum-based drugs is extremely affected by serious systemic toxicities and drug resistance. The affinity of given metallodrugs is to interact with DNA that has been measured as a function of the decrease of guanine oxidation signal on a DNA electrochemical biosensor. In this chapter, the potential activities of metallodrugs to described and representative examples from the most recent families of metal-based compounds are discussed with respect to their possible mode of action and most probable biomolecular targets. We seek to give an overview of the cytotoxic effect of metallo­drugs and also focus on metal complexes of platinum, copper, and zinc with an emphasis on the new strategies used in the development of more and newly designed metal complexes and their biological applications. Keywords:  Metals, Pt-complexes, Cu-complexes, Zn-complexes, DNA, cytotoxicity

*Corresponding author: [email protected] Shahid-ul-Islam, Athar Adil Hashmi and Salman Ahmad Khan (eds.) Advances in Metallodrugs: Preparation and Applications in Medicinal Chemistry, (71–114) © 2020 Scrivener Publishing LLC

71

72  Advances in Metallodrugs

3.1 Introduction Inorganic chemistry is not the “Dead Chemistry” that some people may think. Inorganic compounds were used in the medicinal field for many years. Nowadays, it is well-known that metal ions are essential ingredients in life, just like as organic molecules. In biological systems, several metals play an important role and the field of knowledge concerned with the applications of inorganic chemistry to therapy or diagnosis of a disease is medicinal field. Some metals show many critical functions in humans, and the best examples contain growth retardation which arising from insufficient dietary zinc, pernicious anaemia resulting from iron deficiency, and the heart disease in infants due to copper deficiency. The ability to recognize the treatment of diseases caused by inadequate metalion function constitutes an important aspect of medicinal bioinorganic chemistry [1–3]. The serendipitous discovery of cisplatin by Rosenberg in 1965 unlocked the gate of the unexplored world of metal-based chemotherapeutic agents [4]. These have diverse kinetics and mechanism of action from those of conventional organic drugs. However, the big advantages of metal-based compounds especially transition metals exhibit certain properties including their potential to undergo a redox reaction [5]. Therefore, metals and their redox behaviors are strongly regulated to maintain normal well-being [6–9]. Metals can change the pharmacological properties of organic-based drugs by forming coordination complexes with them. According to Hambley, metal-based pharmaceuticals can be divided into seven different categories depending on the function of the metals and drugs (ligands) moiety such as: (1) the metal complexes are active in its reactive form, (2) metal complexes are active in its inert form, (3) the metals serves as a radiation enhancer, (4) the compounds contains a radioactive metal, (5) the metals or its biotransformation products are active, (6) a ligand is biologically active, and (7) only a fragment of the complex is active. The interaction of drugs and metal-based compounds towards DNA is also an important aspect of pharmacology. DNA is generally the main intracellular target of anti-cancer drugs, so the interaction of the small molecules with DNA can cause DNA damage in cancer cells, blocking the separation of cancer cells and causing the cell death. Metal-based complexes can bind to DNA via both covalent and/or noncovalent interactions. In case of covalent bonding, the labile ligand of the complexes can be displaced by a nitrogen base of DNA such as guanine N7, whereas noncovalent interactions include three different binding modes such as intercalative,

Metallodrugs in Medicinal Inorganic Chemistry  73 electrostatic and groove (surface) binding of metal complexes outside of DNA helix, along a major or minor groove. The interaction between the metal complexes and DNA can cause chemical and conformational modifications and variation of the electrochemical properties of nucleobases [10–12]. Newly, there has been a great interest in the binding of metal complexes with DNA, owing to their possible applications as new cancer therapeutic agents [13–15]. One of the most approaches has been to inhibit the growth of cancer cells is by disrupting the flow of their genetic information. Metal-based drugs have for long been of interest, but the interest of the scientific community for medicinal aspects of metal compounds has rapidly grown in the last few decades [16]. Even in the presence of new chemotherapies, metallodrugs alone or in combination therapy regimen will continue to make up part of the toolkit to fight against cancer [17]. Medicinal inorganic chemistry is a field of increasing prominence as metal-based compounds offer possibilities to design of therapeutic agents which are not readily available to organic compounds. There is a lot of chemical space to explore; furthering our understanding of how metal complexes exert their biological effects and further capabilities they may have to design better drugs for the future of cancer treatment [18, 19]. The uses of metal-based complexes as therapeutic agents, Platinumbased drugs are the mainstay of chemotherapy regimens in the clinic. Nevertheless, the efficacy of platinum drugs is badly affected by serious systemic toxicities and drug resistance, and the pharmacokinetics of most platinum drugs is largely unknown. Copper accumulates in tumors due to the selective permeability of the cancer cell membrane to copper complexes thereby they can act as “artificial nucleases” for the sequence-specific disruption of gene function. Copper is the third most abundant trace element after zinc and iron in the cellular body. It shows the potential synergetic activity with drugs and biological roles in proteins include oxygenation, dioxygen transport, and electron transfer [20]. It is essential for regulated the strength of the blood vessels, skin, urinary tract, and connective tissue in the body [21]. Copper accumulates in tumors due to the selective permeability of the cancer cell membrane to copper complexes thereby they can act as “artificial nucleases” for the sequence-specific disruption of gene function. Recently, it was shown that copper proteins are associated with metabolic changes in cancer cells and most importantly play a significant role in angiogenesis by stimulating proliferation and migration of human endothelial cells, while zinc as an essential element plays a vital role in many physiological and pathological processes. Zinc controls cell proliferation, differentiation, and viability including apoptosis [22–24]. Over the

74  Advances in Metallodrugs past several decades, various antimony complexes were also used for the treatment of leishmaniosis. Medicinal inorganic chemistry as a discipline, however, started to develop after the serendipitous discovery of the anti-­ tumor activity of cisplatin.

3.2 Platinum Anticancer Drugs Over the past years, Platinum complexes are well-known for their antitumor activities. More than thousands of platinum-based complexes have been prepared and tested as potential anticancer agents. A list of Platinum-based metallodrugs, its use, and its mode of action in brief are listed in Table 3.1. One of the best-known and oldest metallodrugs (anticancer-drug) cisplatin, cis-diammine-dichloroplatinum(II) also known as Platinol, Cisplatin discovery as a treatment for cancer and its approval by the US FDA in 1978. It first synthesized by Peyrone in 1844s and in the 1960s, its anticancer properties were discovered by Rosenberg and co-workers [25–27], structure shown in Figure 3.1d, it is widely used that and often referred to as the penicillin of cancer. During entering, inside the cell, one or both of cisplatin’s chlorido ligands are exchanged with water molecules and this aquated platinum (II) species react with nitrogen donors of DNA bases to inhibiting transcription and triggering apoptosis [28]. The researcher has focused on designing the non-classical platinum structures including transplatin, monofunctional, and multinuclear platinum complexes. Non-functional phenanthriplatin, cis-[Pt(NH3)2(phenanthridine)Cl]+ kills cancer cells more effectively than cisplatin because it form monofunctional DNA adducts [29]. In the mode of action, Cisplatin provided the first evidence that Platinum interacts with DNA could be of great importance. Cisplatin forms coordinative DNA adducts such as 1,2-intrastrand crosslinks at d(GpG) sites. It is generally believed that the therapeutic effect of cisplatin is activated by the crosslinking of DNA [30, 31]. Research work related to platinum drugs for anticancer therapy comprised this concept and tried to overcome its various drawbacks mainly lowering the level of cytotoxicity with second-generation platinum drugs like oxaliplatin and carboplatin, etc. One of the metallodrugs of platinum is Satraplatin, Figure 3.2e. The advantage of the Satraplatin has oral availability and can be administered in pill form which is convenient for the patient and reduces health care costs. Triplatin tetranitrate, BBR3464, Figure 3.2a,

Metallodrugs in Medicinal Inorganic Chemistry  75 Table 3.1  A list of Platinum-based metallodrugs, its use, and its mode of action. S. n.

Pt- metallodrugs

Significant comments

01.

Cisplatin

Used for penicillin of cancer. Platinum (II) species react with nitrogen donors of DNA bases to inhibiting transcription and triggering apoptosis.

02.

Oxaliplatin

Oxaliplatin is drugs of platinum-containing in similar design manner are nedaplatin, lobaplatin, and heptaplatin which are currently in clinical trials in the U.S. It appears to reduce serious adverse reactions of the anticancer activity of the platinum and is currently undergoing clinical trials.

03.

Carboplatin

It is six-membered ring reduces its chemical reactivity and possible side effects as well as damage to the ear and the kidneys.

04.

Nedaplatin

Same as Oxaliplatin which appears to reduce serious adverse reactions allowing better exploitation of the anticancer activity.

05.

Lobaplatin

Same as Oxaliplatin, which appears to reduce serious adverse reactions allowing better exploitation of the anticancer activity of the platinum agent.

06.

Heptaplatin

It is also same as Oxaliplatin which appears to reduce serious adverse reactions and a platinum agent is currently undergoing clinical trials.

07.

Picoplatin

Picoplatin is superior and made to overcome cisplatin resistance by adopting different candidate ligands. It is effective in chemotherapy against solid tumors because its kinetic profile of biomolecule interactions is different.

08.

Satraplatin

Satraplatin has oral availability and can be administered in pill form which is convenient for the patient and reduces health care costs. (Continued)

76  Advances in Metallodrugs Table 3.1  A list of Platinum-based metallodrugs, its use, and its mode of action. (Continued) S. n.

Pt-metallodrugs

Significant comments

9.

Polyaminebridged polynuclear complexes

Polyaminebridged complexes (1, 1/t,t) probably has similarities in mode of biological action with cisplatin.

10.

BBR3464

BBR3464 is an unusual trinuclear platinum complex with an overall charge of +4 and in phase II clinical trials. Lung cancer patients did not show a significant response.

11.

TriplatinNC

TriplatinNC is non-coordinatively bound to DNA by a phosphate clamp. It is noteworthy that the cellular uptake of the trinuclear Pt(II) complexes is more efficient than that of cisplatin.

O H3N

O O

H2N

Pt H3N

(a) Nedaplatin

H 3N

II Pt

O

H3N

O

O

(b) Oxaliplatin

O

H3N

Cl

O

H3N

O O O

(c) Carboplatin

II Pt

H3N

O O O

(d) Cisplatin

H3N

II Pt

O

Cl Pt

H3N

O

O H2N

(e) Heptaplatin

H 3N

II Pt

O O O

(f) Lobaplatin

Figure 3.1  The structural formula of the metallodrugs of platinum: (a) Nedaplatin, (b) Oxaliplatin, (c) Carboplatin, (d) Cisplatin, (e) Heptaplatin, and (f) Lobaplatin.

is an unusual trinuclear platinum complex with an overall charge of +4 and in phase II clinical trials, lung cancer patients did not show a significant response to BBR3464 while experiencing toxicity associated side effects such as neutropenia and diarrhoea due to this reason further clinical

Metallodrugs in Medicinal Inorganic Chemistry  77 H3N

H2 N

H2N

NH3

H3N

Pt

Cl

NH3

Pt

Pt

H2N

N H2

4+

Cl

H3N

NH3

(a) BBR3464

H3N +

H2N

H3N

Pt

H2 N

H2N

NH3

H3N

NH3

Pt

H3N H2N

N H2

Pt

8+

+

NH2

NH3

NH3

(b) Triplatin NC

H 3N

Cl

L

X

Pt

N

Cl

L = ammine or amine L1 = N-donar aromatic heterocycles X = chloride or acetate

Pt X

L1

(c) Picoplatin

(d) Trans-platinum complex

O H2 O N Pt H 3N O

Cl

H3N

Cl

Cl

NH2

H2N

Pt n = 2 to 6

NH3

2+

Pt NH3

n H3N

Cl

O (e) Satraplatin

(f) Polyamine-bridged polynuclear complexes, 1,1/t,t

Figure 3.2  The structural formula of the metallodrugs of platinum: (a) BBR3464, (b) TriplatinNC, (c) Picoplatin, (d) Trans-platinum complex, (e) Satraplatin, and (f) Polyamine-bridged polynuclear complexes.

development of BBR3464 was stopped [32]. The interaction of multinuclear platinum(II) drug BBR3464 with DNA by forming so-called phosphate clamps to elicit cell death [33]. Carboplatin (Figure 3.1c) was one of the metallodrugs of platinum reported by Cleare and Hoeschele in 1973 and approved by the FDA in 1989. The chelate effect of the six-membered ring reduces its chemical reactivity and possible side effects as well as damage to the ear and the kidneys [34, 35]. Oxaliplatin (Figure 3.1b) also one of the metallodrugs of platinum drugs was approved by the FDA in 2002. Drugs of platinum-containing in similar design manner are Nedaplatin

78  Advances in Metallodrugs (Figure 3.1a), Lobaplatin (Figure 3.1f), and Heptaplatin (Figure 3.1e) which are currently in clinical trials in the U.S. but are already in clinical use in Japan, China, and South Korea, respectively. In addition, which appears to reduce serious adverse reactions allowing better exploitation of the anticancer activity of the platinum agent is currently undergoing clinical trials [36, 37]. This cross-resistance polyamine bridged complexes (1, 1/t,t) shown in Figure 3.2f is probably have similarities in mode of biological action with cisplatin. An NMR-derived structure of a complementary DNA octamer, d(CATG1CATG)-d(CATG1CATG) (where G1 is a N7-platinated guanine) crosslinked with the Platinum complex (1,1/t,t), [{trans-PtCl(NH3)2}2(µ-NH2(CH2)nNH2)]2+ (n = 4), this structure disrupts the Watson-Crick base pairing. Picoplatin is metallo­ drugs of platinum which is superior and made to overcome cisplatin resistance by adopting different candidate ligands, Figure 3.2c. It is a compound that is effective in chemotherapy against solid tumors because its kinetic profile of biomolecule interactions is different. One of the main factors is an adaptive increase in the rate of intracellular detoxification by glutathione and metallothioneins [38]. Picoplatin is a sterically hindered Pt(II) complex towards thiol-containing compounds and forms DNA adducts similar to those of cisplatin. Picoplatin is now a chemotherapeutic agent in clinical development and improved the toxicity profile relative to existing platinum-based cancer therapies and demonstrated clinical efficacy in platinum-refractory small cell lung cancer [39]. Successive testing of Pt(II) metallodrugs compounds of Pt has had the general formula of cis-[Pt L2 X2] (where L represented ammine or amine, and X represented a leaving group such as halide or carboxylate). In trans-Pt(II) complexes, a similar approach has been made by Farrell et al. The trans-Pt(II) complexes are the stereo-isomer of cisplatin that was found to be therapeutically inactive in the 1960s. Therefore, in the classic structure-activity relationship, it has been usually noted that antitumoractive Pt(II) complexes possess two kinetically inert Pt−N bonds and two relatively labile Pt-X (X = chloride or carboxylate group) bonds with a cis geometry and substitution of a range of amines for the ammine ligand of transplatin yields trans-Pt(II) complexes with significant cytotoxicity. The interactions of cisplatin with DNA yield interstrand crosslinks with the adjacent guanines of CG/GC base pairs [40]. Monofunctional Pt(II) complexes, PtL3X, are usually found to be non-cytotoxic or only moderately cytotoxic. Mononuclear Pt(II) complex, [PtCl(en)(ACRAMTU-S)]2+, en = ethylenediamine, ACRAMTU = 1-[2-(acridine-9-ylamino)-ethyl]-1,3dimethylthiourea; which exhibit cytotoxicity at micromolar to nanomolar concentrations in a wide range of solid tumor cell lines [41]. Derivatives

Metallodrugs in Medicinal Inorganic Chemistry  79 of Figure 3.3C that include [PtCl(en)(N-(acridin-9-yl)-Nµ-methlethane1,2-diamine)]2+ (Figure 3.3A) and [PtCl(NH3)2)(N- (acridin-9-yl)-Nµmethlethane-1,2-diamine)]2+ (Figure 3.3E) show remarkably high cytotoxicity in H460 NSCLC cells and were found to inhibit the rate of tumor growth by 40% in a H460 mouse xenograft study [42]. This Phenanthriplatin or cis-[Pt(NH3)2-(phenanthridine)Cl]NO3 is a new drug candidate of the metallodrugs of platinum, Figure 3.3F. It belongs to a family of platinum(II) Phenanthriplatin that was discovered by Professor Stephen J. Lippard at Massachusetts Institute of Technology and is currently being developed by Blend Therapeutics for its potential use in human cancer therapy [43]. Phenanthriplatin kills cancer cells more effectively than cisplatin by

2+

H3N

NH2

H2N Pt Cl

HN

OH

N N H

Pt NH3

H3N

3A

3B

2+

Pt Cl

Pt

N

NH S

N

N

N H

OH

N

Pt NH3

H 3N

3D

3C

+ NH3

N Pt Cl

NH3

3F- Phenanthriplatin

2+

NH3

H 3N

NH2

H2N

2+

Pt

N

NH

N

NH3

O – N+ – O O

2+

NH3

H3N Pt Cl

NH HN

N N H

3E

Figure 3.3  Structural formula of the series of Pt-acridine complexes (3C, 3A, 3E), Azolato-bridged dinuclear platinum(II) complexes 3B and 3D and Phenanthriplatin 3F.

80  Advances in Metallodrugs forming monofunctional DNA adducts [44]. A large number of azolatobridged dinuclear Pt(II) complexes like [{cis-Pt(NH3)2}2(µ-OH)(µ-pyrazolato)]2+ (Figure 3.3B) and [{cis-Pt(NH3)2}2(µ-OH)(µ- 1,2,3-triazolato)]2+ (Figure 3.3D) have been developed as a new class of Platinum containing metallodrugs which is capable of crosslinking two adjacent nucleobases without causing kinks in the double helix and this effect could be sufficient to trigger the desired cytotoxic actions and allow remarkable in vitro cytotoxicity in several human tumor cell lines [45]. Therefore, the basic skeleton of the azolato-bridged dinuclear Pt(II) complexes will be useful in developing the next-generation anticancer metallodrugs of antitumor drugs.

3.2.1 Nucleophilic Displacement Reactions in Complexes of Platinum Square-planar complexes of platinum undergo predominantly bimolecular nucleophilic displacement reactions in contrast to the generally dissociative reactions exhibited by the octahedral complex. Cis-[Pt(NH3)C12], for example, is injected as a neutral molecule in physiologic saline solution. This neutral species may undergo limited hydrolysis in the extracellular fluid (0.1M Chloride) [46]. Within the cell, a lower chloride ion concentration obtains (0.004 M Chloride); thus, more extensive hydrolysis may occur in the stepwise reaction mechanism. The rate constants for the successive reactions for cis-[Pt (NH3)2 Cl2] are represented as k1(H2O) = 2.5 × 10−5 sec−l, k2 (H2O) = 3.3 × 10−5 sec−1; at 20ºC; thus, these reactions have half-times of the order of 6–8 h (Figures 3.3A, C, and D).

3.2.2 Mode of the Interaction of Cisplatin Species With Nitrogen Donors of DNA Strand Metallodrugs of Platinum like cisplatin crosslink interact with Comple­ mentary octamer present at DNA, Binding of cisplatin to DNA is irreversible, and structurally different adducts are formed by either intrastrand crosslinking of two nucleases of single DNA strand, interstrand crosslinking of two different strands of one DNA molecule. During crosslinking with DNA strands, a chelate formation through N and O atoms of one guanine and DNA protein crosslinks shown in Scheme 3.1 and Scheme 3.2, respectively. Trinuclear complex of Pt(II) is noncoordinating complex, [{transPt(NH3)2(NH2(CH2)6(NH3 +)}2-µ-{trans-Pt(NH3)2(NH2(CH2)6 NH2)2}]8+

Metallodrugs in Medicinal Inorganic Chemistry  81 also known as TriplatinNC (Figure 3.2b). TriplatinNC is noncoordinatively bound to DNA by a phosphate clamp in which the square planar tetra(m)mine Pt(II) coordination sphere unit forms bidentate complexes with phosphate oxygen atoms shown in Scheme 3.3. The detailed TriplatinNC non-coordinative interaction modes with DNA dodecamer Cis - [Pt(NH3)2(OH)2]0 pKa = 7.3 –H+ +H+ +H2O

Cis - [Pt(NH3)2(OH)2]0

+Cl–

Cis - [Pt(NH3)2(H2O) (OH)]+

–H+ +H+ Cis - [Pt(NH3)2 Cl2]0

+H2O +Cl–

pKa = 5.6 –H+ +H+ +H2O

Cis - [Pt(NH3)2(H2O) Cl]+

Cis - [Pt(NH3)2(H2O)2 ]2+

+Cl–

Scheme 3.1  Displacement reactions involve in square-planar metallodrugs complexes of platinum. H3N

NH3 Pt G G

G

H 3N

H3N

H3N

Pt NH3

G

Pt

G

Pt

NH3

G G

G

NH3

Scheme 3.2  Metallodrugs Cisplatin species react with nitrogen donors of DNA bases to inhibiting transcription and triggering apoptosis.

O HN

−

O

P

O

NH2 Pt 2+

Phosphate of DNA

O

H2N

NH

Scheme 3.3  Schematic representation of a phosphate clamp, in which the square planar Pt (II) coordination sphere unit forms bidentate complexes with phosphate oxygen atoms.

82  Advances in Metallodrugs [d(CGCGAATTCGCG)]2 were exposed by X-ray crystal analysis [47]. It is noteworthy that the cellular uptake of the trinuclear Pt(II) complexes is more efficient than that of cisplatin.

3.2.3 Systemic Toxicity of Cisplatin Despite its importance in clinics, it leads to severe side effects of cisplatin in systemic toxicity like severe nausea, vomiting, loss of hearing, and kidney damage when used for chemotherapy. These adverse effects mean that many patients are treated with less than optimal doses, causing tumors such as ovarian cancers to rapidly develop resistance [28]. The long-term side effects of platinum therapy are significant and some patients have to undergo dialysis, kidney transplant, or take a cocktail of medicines to support other systems in the body.

3.3 Copper-Based Anticancer Complexes 3.3.1 Copper is Essential for Health and Nutrition Copper is the most studied metal in all transition metal ions and very essential for all living things. As a naturally-occurring element, copper exists everywhere in the world around us. Life has evolved in this natural presence, and humans have developed built-in mechanisms to manage intake levels. Copper is not formed in the body and must be obtained from food and drinking water each day as part of a balanced diet and, occasionally, through the use of dietary supplements. Dietary copper is important to doctors and nutritionists [48–50]. Our digestive systems assimilate the amount necessary for good health through a system of uptake called homeostasis. Excess copper is excreted [51]: Copper is Essential for: • Maintenance of healthy skin and connective tissue • Growth of new blood vessels • Generation and storage of energy in the “power plants” of our cells, the mitochondria • Formation of the cells of our immune system (white blood cells) • Wound healing

Metallodrugs in Medicinal Inorganic Chemistry  83 • Brain development during foetal and postnatal growth, and maintenance of brain health throughout life, including effective anti-oxidative defence • Efficient communication between nerve cells • Structural integrity and function of heart and blood vessels • Proper structure and function of circulating blood cells • Maintenance of a healthy and effective immune response

3.3.2 Healthcare Applications of Copper With so many HAIs being reported, not just in the United States, but around the world, medical care facilities have implemented copper in their everyday operations. In order to take advantage of its special properties, copper is used in a number of health applications such as [52]: Copper bed railings are a major point of contact for patients. Those with infections frequently touch the surface of bed rails in order to help them get up and down. Copper will prevent bacteria from remaining on the surface. Door handles and knobs are everywhere in a hospital and can be touched by patients from all parts of the facility. Using copper handles can help prevent. I.V. poles are moved around by both patients and staff. Using copper alloy poles can reduce the risk of infection to everyone since a virus will not survive on the surface of the pole. Faucets can also be a haven for surface microorganisms, as well as those that survive in droplets. Copper alloy faucets create a hostile surface for these microbes, reducing the risk of infection through secondary or droplet contact. Copper has many useful properties and, for the healthcare industry, its antimicrobial properties are the most valuable. Healthcare providers around the world are now using copper in various applications which allow better care and reduced risk of infection for everyone

3.3.3 Copper and Human Health Disorders There are some rare and inherited diseases: Wilson’s disease, Menkes’ Disease, and Cancer Disease which are genetic in origin and which are brought about by disturbances to the management of the bodily stores of copper [53–55].

84  Advances in Metallodrugs Copper is very crucial for the normal formation of the brain and nervous system. It also plays a significant role in making nuerotransmitters, the chemical messengers that facilitate communication between nerve cells, and the movement of electrical impulses along nerves.

BLOOD VESSELS AND HEART: Copper helps to sustain the clasticity of blood vessels, which allows maintenance of proper blood pressure. The aorta-the main artery that runs from the heart, and the largest in the human body-cannot function fully if its elastic framework is weakened. Since copper is needed for the healthy muscle tone and function, it also plays a vital role in the heart.

THE BONE: Copper play an important role in collagen formation and is crucial for bone formation, health and repair. Collagen is the primary factor for the rigidity, mechanical strength and competence of bone. In fact, animal studies show that bone fractures, skeletal abnormalities and osteoporosis occur with copper deficiency

Copper

White blood cells Foreign material

THE SKIN: Copper plays an vital role in collagen formation, formation, a connective tissue in the skin. Collagen is the most prevalent prevalent found in human skin and is important in maintaining our appearance-supply, healthy appearing, wrinkle-free skin-on our faces and all other areas.

The immune system: Copper is necessary for the maintenance of a healthy immune system to ward off germs and diseases. A strong and aggressive supply of germ-fighting soldiers, including white blood cells (engulf foreign material), anti-bodies (protein molecules), B lymphocytes (produce antibodies) and T lymphocytes (immune cells), keep the body healthy and disease-free.

3.3.3.1 Wilson’s Disease (WD) The basic characteristic of Wilson’s disease, which was first described in 1912 [56], is a massive accumulation of copper in the liver and brain due to the inability of Wilson’s Disease patients to transport copper out of the affected tissues via the blood protein ceruloplasmin or via 9 biliary excretion [57]. Ultimately, the accumulation of the copper leads to nervous disorders (mild tremors), and to pathological lesions especially in the liver. In extreme cases, the light brown circles referred to as Kayser-Fleischer rings surrounding the iris are seen, which are caused by the deposition of

Metallodrugs in Medicinal Inorganic Chemistry  85 copper salts in the cornea [58]. While much is known about the nature of Wilson’s disease which affects an estimated 1 in 160,000 people, the cause is as yet unknown. Patients suffering from Wilson’s disease are treated with such drugs as the copper chelating agents penicillamine (introduced in the 1960s), that removes or modulates copper distribution [59, 60].

3.3.3.2 Menkes’ Disease Symptoms of Menkes’ Disease, an X-linked genetic disease of males, appear before the third month and usually terminate the life of the child before the fifth of the sixth year. No cure is yet known and attempts to control the course of the disease with copper, therapy have proved disappointing despite correction of low liver and serum copper levels [61]. Described first in 1962s [62], the disease causes retarded growth with progressive neurological and vascular disturbances, pallid skin, and brittle hair, often referred to as steely or kinky. The cerebral function is grossly affected and there is progressive brain degeneration [63]. A general impairment in copper metabolism was postulated as the major cause of the disease following the finding that Menkes’ patients have abnormally low amounts of copper in liver and serum [64]. Unfortunately, oral administration of copper to Menkes’ patients is very poorly absorbed, with the copper tending to accumulate within the mucosal cells [65]. Intravenous copper therapy is normally only partially successful in treating the disorder [66]. Further parallels between copper deficiency and Menkes’ disease have been revealed including similar if not identical connective tissue pathology and imperfections of the arterial walls. While it is agreed that the intracellular metabolism of copper in Menkes’ cells is quite different from that in normal cells, there is as yet no consensus on the mechanism involved. The main difference in pathology between genetic and nutritional copper deficiency relates to iron; while anaemia is often observed in nutritional copper deficiency in many species, it does not occur in the genetic copper deficiency diseases.

3.3.4 Role of Copper Complexes as Potential Therapeutic Agents Copper forms a rich variety of coordination complexes with oxidation states Cu2+ and Cu+, and very few examples of Cu3+ compounds are reported [67]. The design, synthesis, and development of copper-based complexes as anticancer agents were reported by several reviews over the last decade [68]. Nowadays, copper-based complexes are considered the best alternative

86  Advances in Metallodrugs to cisplatin drugs and are well known to have great importance in cancer chemotherapy. Copper atoms also exhibit numerous biological applications and synergetic activities with drugs. In many cases, it has been found that copper complexes containing N-donor heterocyclic ligands, such as 2,2’-bipyridine (bpy), 2,2’-dipyridylamine (bipyam), 1,10-phenanthroline (phen), bathophenanthroline (Bphen), etc., possess more biological affinity than the parent drugs [69]. The coordination of metal ions with quinolone drugs is of great interest from the biological and pharmaceutical point of view because it increases the binding affinity towards DNA and proteins as well as the nuclease activity towards genomic, plasmid, and internucleosomal DNA [70]. Many Cu(II) complexes of quinolones derivatives, Schiff bases and NSAIDs showing enhanced anti-inflammatory and antiulcerogenic activity, as well as reduced gastrointestinal toxicity compared to the uncomplexed drug, have been prepared and structurally characterized [71, 72]. The different biological activity of these complexes compared to one of the widely used platinum anticancer drugs cisplatin indicates different mechanism(s) of action, which have not been yet resolved. It is likely that copper complexes interact with enzymes and inhibit vital cell functions, rather than interact with DNA and induce crosslinks. This chapter includes representative examples of copper-based complexes according to ligand similarity that has been tested for their anticancer activity in about the last 10 years. It has been established that the copper ion chelation plays a definite role in the biological activity of most of the selected organic ligands, enhancing the anti-tumor activity of the metal-free substances.

3.3.4.1 Thiosemicarbazones-Based Complexes Thiosemicarbazones (TSCs) belongs to a class of compounds of medicinal interest whose anticancer activity has been reported as early as the 1960s [73] and their synthesis is still in progress [74–77]. TSCs exit as thione-thiol tautomers which can bind to a metal center in the thione as well as in the anionic forms as seen in Figure 3.4.

R1 R2

N1

H

R3

N2

N3

R1 R4

S

(a; thione)

Figure 3.4  TSCs frameworks.

R2

N

H

R3

N

N SH

(b; thiol)

R1 R4

R2

N

H

R3

N

N S–

(c; anion)

R4

Metallodrugs in Medicinal Inorganic Chemistry  87 A number of bonding modes have been observed for TSCs in their neutral or anionic forms, and depending on the substituents, they can behave as N,S bidentate, N,S,D (D = N,O) tridentate, or N2,S,D (D = O, S) tetradentate ligands. Binary Cu(II) complexes with a variety of aromatic molecules coordinated through N, S, or O donor atoms have been synthesized and tested for biological activity. Some of them are presented in Figure 3.5. The copper(II) complexes [Cu(L5a)Cl]·DMSO (5a1) and [Cu(L5a)Br]·DMSO (5a2) (HL5a = pyridine-2-carbaldehyde-TSC) (Figure 3.5a) showed a distorted bipyramidal coordination polyhedron of the complexing ion in which the TSC sulfur atom served as a bridge and occupied the copper fifth coordination site. In the crystal structure, the [Cu(L5a)]-NO3·DMSO (5a3) complex (Figure 3.5b) formed polymer chains in which the copper atom of one complex coordinated with the thiocarbamide nitrogen atom of the neighboring complex. 5a1, 5a2, and 5a3 complexes at a concentration of 10−5 mol/L selectively inhibited the growth of 60%−90% of HL60 cells [78]. Interaction of 5a3 and [Cu(L5b)(NO3)] [HL5b = pyridine-2-carbaldehyde 4-N-methyl-TSC (Figure 3.5c)] with [poly(dA-dT)]2, [poly(dG-dC)]2, and calf thymus DNA (CT-DNA) has been probed and they exhibited toxicity against V-79 cells in the low micromolar range (IC50 = 6.14 and 2.43 μM, respectively) [79].

(a) N

NH2

Cu

S

X S H2N N

N

N DMSO

.2DMSO Cu

NH2

N

(b)

N

N

N

X

N

[Cu(L5a)X]2.2DMSO 5a1: X = Cl 5a2: X = Br

S Cu

NO3 NH2

N N S N

Cu

{[Cu(L5a)] NO3.DMSO} 5a3

NO3

DMSO

H3C

(c)

NO3 NH

N N S N

Cu

[Cu(L5b)H2O] NO3 5b1

H2O

Figure 3.5  Binuclear formylpyridine-TSC copper(II) complexes of (a and b) HL5a = pyridine2-carbaldehyde-TSC or formylpyridine-TSC; (c) HL5b = pyridine-2-carbaldehyde 4-N-methyl-TSC.

88  Advances in Metallodrugs

3.3.4.2 Quinolones-Based Copper Complexes Antibacterial quinolone and fluoroquinolone drugs, such as nalidixic acid (nal), enoxacin, moxifloxacin (mof), norfloxacin, levofloxacin (lef), and ciprofloxacin (cp), have been extensively used together with N-donor heterocyclic ligands such as 2 2’-bipyridine, 2 2’-dipyridylamine, 1–10 phenanthroline, etc., for synthesis of ternary copper complexes, but other examples of copper(II)−quinolones complexes have been reported where the quinolone acted as a bidentate ligand coordinating the metal through carbonyl oxygen and carboxylate oxygens. In a series of five neutral metal complexes [Fe(mof)3], [V(O)(mof)2], [Cu(mof)2(H2O)2], [Ni(mof)2(H2O)2], and [Mn(mof)2], the copper derivative showed the strongest DNA binding via intercalation and the highest in vitro antiproliferative activity against A549 cancer cells (IC50 5.4 μg/mL) [80]. Square pyramidal, ternary Cu(II) complexes 6a–f were prepared with ciprofloxacin and bidentate N, S-donor ligands (L6a–f; Figure 3.6). By changing the electronic properties of the intercalative ligand, a variation on the DNA interaction ability and on the cytotoxicity of the compounds was observed [81].

3.3.4.3 Naphthoquinones The 1,2- and 1,4-naphthoquinone moiety are usually encountered in natural products, and their derivatives show a wide range of biological applications including anticancer activities [82, 83]. Such properties are due to the interference of quinones in the electron transport chain by electron reduction processes, generating semiquinone radical (Q•−) and hydroquinone anion (Q2−) [82, 83]. Incorporation of an azo group into 2-hydroxy-1,4-naphthoquinone has led to promising anticancer agents HN N

F

6a: R = F 6b: R = H 6c: R = Cl 6d: R = Br 6e: R = CH3 6f: R = OCH3

Cl N

O O O

Cl

S Cu N

R

Figure 3.6  Structure of copper complexes 6a–f containing ciprofloxacin.

Metallodrugs in Medicinal Inorganic Chemistry  89 R

O

N

N O

O

O

Cu

O N

N

O

R

7a: R = 4-OMe 7b: R = 4-N=NC6H5 7c: R = 4-Cl 7d: R = 4-I 7e: R = 3-I 7f: R = 2-I 7g: R = 4-COOH 7h: R = 3-COOH 7i: R = 4-CN 7j: R = 3-CN 7k: R = 4-NO2 7l: R = 3-NO2 7m: R = 2-NO2

Figure 3.7  Structure of Cu−naphthoquinone complexes a–m.

in which metal complexation with copper(II) resulted in increased cytotoxicity [84]. The relative stability of the tautomeric forms of 3-(2-R-phenylhydrazono)-naphthalene-1,2,4-trione derivatives is a function of the substituents (Figure 3.7) [85]. Some of these ligands and related [Cu(La−m)2] complexes 7a–m (Figure 3.7) have shown in vitro antitumor activity against several cancer cell lines (SF295, HCT-8, and MDA-MB-435). Free hydrazono HL7f and complex 7m exhibited higher growth inhibition of HCT-8 cells (96.03%) than the positive control doxorubicin (dox) (91.67%). The results indicated that the presence of −NO2 and −I groups were relevant for the antitumor activity. Formation of an organic free radical observed in the EPR spectrum of [Cu(L7m)2] might be responsible for the cytotoxic activity [86, 87].

3.4 Zinc Anticancer Complexes Zinc is the second most trace mineral in the body. The trace elements were relatively slow to be recognized as essential elements, whereas iron (Fe) was the first. In the 17th century, anaemia was verified to be caused by an iron deficiency and often was cured by supplementing the diet with extracts of rusty nails. In the 19th century, trace amounts of iodine were found to eliminate goitre that is an enlarged thyroid gland. Copper (Cu) was shown to be important for humans in 1928, and manganese, zinc (Zn), and cobalt and elemental composition present in the human being shown in (Table 3.2). After taking these accounts, the Zn, Cu, and platinum (Pt) as a metallodrug have been used in medicinal chemistry.

90  Advances in Metallodrugs Table 3.2  Approximate elemental composition of a typical 70-kg human being. Trace elements (mg)

Trace elements (mg)

Zinc

1,750

Iron

5,000

Barium

21

Rubidium

360

Silicon

3,000

Copper

280

Molybdenum

14

Strontium

280

Bromine

140

Tin

140

Manganese

70

Iodine

70

Aluminium

35

Boron

14

Arsenic

~3

Chromium

~3

Cobalt

~3

Nickel

~3

Vanadium

~2

Selenium

~2

Lead

35

Lithium

~2

3.4.1 Biologically Importance of Zinc Zinc is a trace metallic element with atomic number 30, atomic weight 65.4, essential for an important role in all living being. It is the 24th most abundant trace element and diamagnetic properties. Because of its nature as a transitional element in the periodic table, zinc possesses certain chemical properties that make it especially useful and important in biological systems have been used as a micronutrient. Additionally, zinc is able to constitute strong but readily exchangeable and flexible, complexes with organic molecules, thereby enabling it to modify the three-dimensional structure of nucleic acids, specific proteins, and cellular membranes. In 1961, the need for zinc for the human body was recognized. The total body zinc content of adult human’s ranges from about 1.5 to 2.5 g, most of which are found intracellularly, primarily in muscle, bone, liver, and other organs [88, 89]. The remaining zinc comprises the so-called rapidly exchangeable pool of zinc, which is thought to be particularly important for maintaining zinc-dependent functions of human biological systems such as: • Zinc fingers in DNA • Zinc is required for the synthesis of DNA

Metallodrugs in Medicinal Inorganic Chemistry  91 • • • • • • •

Normal growth Gene expression Gene regulation Cell division Immunity Protein synthesis DNA synthesis

The rapidly exchangeable zinc can move into and out of the plasma compartment within a period of about 3 days. In spite of the proven benefits of adequate zinc nutrition, approximately 2 billion people still remain at risk of zinc deficiency. Zinc is found as a component of more than 300 enzymes and hormones and plays a crucial part in the health of our skin, teeth, bones, hair, nails, muscles, nerves, and brain function as well as it is essential for growth. Zinc controls the enzymes that operate and renew the cells in our bodies. The formation of DNA, the basis of all life on our planet, would not be possible without zinc [90]. Zinc deficiency is an important public health problem, affecting a large number of women and children in India and worldwide. Zinc deficiency is the fifth leading risk factor for disease in the developing world. In a recent survey by WHO, zinc deficiency is found in most of the Indian population and zinc supplement is used commonly to enhance wound healing and treatment of pneumonia [91]. The element is important in maintaining the healthy growth of the human body, especially for infants and young children [92]. Zinc as an essential element for animals [93], fungi [94], including humans being, [95] bacteria, and plants. The first enzyme to be shown to need zinc for activity in 1939 [96], much has been learned about zinc in biological systems, ranging from effects on the whole organismal level, over the identification of important zinc-binding proteins, down to structural, thermodynamic, and kinetic details of zinc-protein interactions [97]. Zinc deficiency affects up to two billion people worldwide. Its multiple systemic effects include growth retardation, weight loss, infertility, mental, and emotional disorders, impaired immune function, skin lesions and hair loss. The most salient recognition of the importance of zinc in human health was delivered by the 2008 Copenhagen Consensus conference, which ranked supplying zinc and vitamin A to over 100 million malnourished children as their highest priority solution to advance global welfare [98]. Besides the drastic consequences of severe zinc deficiency and their alleviation by zinc supplementation [99], the more subtle impacts of zinc homeostasis on ageing, [100] neurodegenerative diseases [101], cancer [102], the immune system [103], and energy metabolism

92  Advances in Metallodrugs [104] are active study areas. Zinc plays ubiquitous biological roles in humans and interacts with a wide range of organic ligands. Stored in specific synaptic vesicles of glutamatergic neurons of the brain, it also helps in modulating brain excitability and is known to play a critical role in synaptic plasticity, cognitive functions, and learning. In the process of DNA replication and transcription inside the cell, zinc finger plays an important role and form part of several transcription factors. Those transcription factors are proteins that help in the recognition of DNA base sequences. Nine to ten Zn (II) ions help in the formation of zinc finger. Demonstration of the benefits and physiological role of zinc in the human body and its deficiency-related impairments in body functions. Zinc fingers to accurately bind to the sequences of DNA. Zinc binds transferrin which in plasma is used to transport iron. The zinc is essential for catalytic properties of many enzyme systems and intracellular signalling. Zinc is associated with more than 50 distinct metalloenzymes, which have a diverse range of functions, including the synthesis of nucleic acids and specific proteins, such as hormones and their receptors. For these reasons, zinc plays a central role in cellular growth, differentiation, and metabolism. Of further interest is zinc’s absence of redox properties, which allows it to be transported in biological systems without inducing oxidant damage, as can occur with other trace elements such as iron and copper.

3.4.2 Schiff Base Chemistry A Schiff base (SB), considered as a condensation product, is typically prepared by the most familiar and classical method discovered by Hugo Schiff, involving the condensation of an aldehyde or a ketone (carbonyl compound) with an amine under diverse reaction conditions and in different solvents [105]. An SB is a nitrogen cognate of an aldehyde or ketone in which the carbonyl group (C=O) has been supplanted by an imine or azomethine (−C=N−) group. The carbonyl group of the aldehydes gives aldimines while ketones give ketoimines.

3.4.2.1 Schiff Base and Their Metal Complexes Herein, we are familiar with different types of Schiff base such as salentype, hydrazine-type, semi-carbazone–type, and thiosemicarbazone. All these types of have a unique and incogitable role in medicine and coordination chemistry.

Metallodrugs in Medicinal Inorganic Chemistry  93 • Salen-type ligand Salen-type ligand has been their preparation from salicylaldehyde, ethylenediamine, and their derivatives. As a special Schiff base, the salen ligand and its derivatives are condensation products with salicylaldehyde and ethylenediamine [106]. The ligand backbone of salen and the coordinated metal ion can be smoothly varied which makes these ligand systems extensively useful in various industrial and biological applications [107]. A wide range of salen-type compounds are present and, depending on the bonding modes, they feature two covalent and two-coordinate covalent bonds occupying equatorial positions, thus behaving as [O, N, N, O] tetradentate ligands with two axial sites occupied by ancillary ligands. • Hydrazone-type ligands Another important class of hydrazone-type ligands [R1R2C=NNHR3] which are a condensation product of carbonyl compound with hydrazine/ hydrazide and their derivatives. Hydrazones and hydrazine derivatives play a vital role in enhancing the selectivity and lethality profile of certain drug candidates [108]. • Thiosemicarbazone/semi-carbazone ligands Thiosemicarbazones/semicarbazones-type ligand made their first appearance in the literature and proved themselves as excellent chelating agents [109]. Thiosemicarbazone/semicarbazone Schiff base represents a family of compounds that are derived via the condensation reaction of a carbonyl compound (aldehyde or ketone) with thiosemicarbazide or semicarbazide. It has the superior chelating ability and extensive lipophilic nature are of outstanding properties. From the literature, studies supported the reality that a wide range of transition metal complexes of these typical Schiff base ligands has been prepared. Thiosemicarbazones/semicarbazones exhibit a diversity of bonding modes due to the occurrence of different donating sites. These types of ligands generally have two donating sites (N, O or S) and hence act as bidentate, but they can behave as tridentate (N, X=N, S, O) ligands also.

3.4.3 Zinc-Based Complexes In recent years, the number of people suffering from cancer and multidrugresistant infections has sharply increased. Although cancer is considered the leading cause of death worldwide. Development of small molecules as anticancer and anti-microbial agents has great potential and a plethora

94  Advances in Metallodrugs

Br

N

O

N

Zn N

N

N N O

O

Zn

O

Cl N

Complex 1

Complex 2

Figure 3.8  Molecular structures of zinc complexes displaying promising better antimicrobial activity.

of drugs are already available to combat these diseases [110]. A successful strategy in anticancer chemotherapy has been the use of metallodrugs and this strategy has the potential to be used for treating multidrugresistant infections more efficiently. As a class of molecules, Schiff bases have been the topic of considerable interest, owing to their versatile metal chelating properties, inherent biological activities, and flexibility to modify the structure to fine tune it for a particular biological application. Schiff base–based metallodrugs are being researched to develop new anticancer and anti-microbial chemotherapies and because both anticancer and antimicrobial targets are different, heterocyclic Schiff bases can be structurally modified to achieve the desired molecule, targeting a particular disease [111, 112]. In this review, we collect the most recent and relevant literature concerning the synthesis of heterocyclic Schiff base metal complexes as anticancer and anti-microbial agents and discuss the potential and future of this class of metallodrugs as either anticancer or antimicrobial agents. According to World Cancer Report 2015, nearly 14.1 million new cases of cancer occurred globally resulting in 8.8 million deaths. The complexes 1 and 2 (Figure 3.8) show the best antibacterial activity with biologically active compounds. Some other Zn-based complexes that exhibit good biological activity are given in Table 3.3.

3.4.4 Top Food Sources of Zinc High-protein foods contain the largest amounts of naturally occurring zinc. Here, are the main top 12 food sources of zinc [113, 114], although keep in mind the absorption rate of zinc is greatest from foods that don’t

2

O

O

N

Zn

N

N

N

N

O

O

N

Zn

N

Zn

N

N

N

N

N

N

2+

.(ClO4)2

Author Dalton Trans., 2013, 42, 5932–5940

Journal

(Continued)

S. Anbu, S. Kamalraj, Inorga. Chem., 2012, 51, B. Varghese, 5580–5592 J. Muthumary, M. Kandaswamy

Zn (II) complexes of S. Liu, W. Cao, L.Li, C. Fan, bis-benzimidazole T. Chen derivatives

Drugs

Acts as a potent anticancer Oxyimine-based agent via inducing macrocyclic phenotypical changes, dinuclear Zn (II) growing membrane complexes permeability, and activation of caspase-3 and -9, which is consistent with the induction of mainly apoptotic cell death

Induced dependent apoptosis in cancer cells by triggering DNA damage thereby act as chemotherapeutic agents used for human cancers

1

N

Function

SN Complex

Table 3.3  Zn-based complexes that exhibit biological activity.

Metallodrugs in Medicinal Inorganic Chemistry  95

4

R

N H

N

Zn

S

N

N

N

R = Me, Et, Ph

S

N

N

N H

N

R

Cl

Cl

Zn N

Zn (II) complex dinucleating polypyridyl ligands

Drugs

Zn (II) bis Shows cytotoxicity comparable to that of (thiosemicar­ cisplatin within a range bazanato of human cancer cell complexes) lines. Their intrinsic fluorescence has permitted the cellular distribution to be imaged, signifying that their activity is associated with disruption of the mitochondria although there is also much slower uptake in the nucleus

Exhibit important cytotoxic activity for the HeLa cell lines and inhibit cell cycle progression through the G1 Phase

3

N Zn Cl Cl N

Function

SN Complex

Table 3.3  Zn-based complexes that exhibit biological activity. (Continued)

S. I. Pascu, P. A. Waghorn, T. D. Conry, B. Lin, H. M. Betts, J. R. Dilworth, R. B. Sim, G. C. Churchill, F. I. Aigbirhio, J. E. Warren

C. Y. Gao, X. Qiao, Z. Y. Ma, Z. G. Wang, J. Lu, J. L. Tian, J. Y. Xu, S. P. Yan

Author

Dalton Trans., 2008, 2107– 2110

Dalton Trans., 2012, 41, 12220–12232

Journal

96  Advances in Metallodrugs

Metallodrugs in Medicinal Inorganic Chemistry  97

Lamb - 3 ounces: 2.9 milligrams

Grass-fed beef 3 ounces: 2.6 milligrams

Pumpkin seeds - ¼ cup: 1.6 milligrams Chickpeas - 1 cup cooked: 2.5 milligrams Yogurt 6 ounces: 1 milligrams

Cashews - ¼ cup: found 1.9 milligrams

Cocoa powder 1 tablespoon: 0.3 milligrams

Chicken - 3 ounces: 1 milligrams

Salmon - 3 ounces: 0.5 milligrams

Turkey - 3 ounces: 1 milligrams Eggs -1 large: 0.6 milligrams

Mushrooms - 1 cup: 0.6 milligrams

Figure 3.9  Schematic representation of top food sources of zinc.

contain any nutrients, which are usually animal-based as opposed to plantbased shown in Figure 3.9.

3.4.5 Role of Zinc in Human Body Zinc has an enormous role and significance such as essential mineral and has vast biological and public health importance. The zinc cation has key roles in enzymatic activity, structural organization, and functional regulation due to its strong Lewis acidity and other unusual chemical properties, including rapid ligand exchange, flexible coordination, and lack of redox chemistry under physiological conditions. However, the physical properties of Zn (II) make most spectroscopic techniques and methodologies employed for probing other biological metals inapplicable to the characterization of zinc enzymes. Synthetic zinc models have attracted much attention because they are more amenable to structural, spectroscopic, and mechanistic studies that could provide information regarding zinc chemistry and its coordination environment. Zinc is distributed widely in plant and animal tissues and occurs in all living cells. It functions as a cofactor and is a constituent of many enzymes like lactate dehydrogenase, alcohol dehydrogenase, glutamic

98  Advances in Metallodrugs Zinc excess lethergy local neuronal defects

Respiratory disorder often take zinc smoke Mental fume fever Diarrhea Nausia/vomiting Epigastric pain

Zinc deficiency Brain

Mental lethergy neurosensor disorder decrease nerve conduction

Brain

trac atory Respir

t

Thym us

Thymic antrophy

ct

tra inal test n i o tr Gas

Skin

Re

te sta Pro

pro

du

Skin lesions

cti

ve s

yst e

m

Elevated risk of pain cancer

Infertility acrodermatitis Less wound healing

Figure 3.10  Schematic representation of zinc in the human body.

dehydrogenase, alkaline phosphatase, carbonic anhydrase, carboxypeptidase, superoxide dismutase, retinene reductase, and DNA and RNA polymerase. Zn-dependent enzymes are involved in macronutrient metabolism and cell replication (Figure 3.10).

3.4.6 Zinc as a Health Benefit The human body contains 2–3 g (2,000–3,000 mg) of Zn and nearly 90% is found in muscles and bones [115]. The nearby organs include prostate, liver, the gastrointestinal tract; kidney, skin, lung, adrenals, brain, heart, eyes, and pancreas contain estimable concentrations of Zn. Blood tests for Zn deficiency are inaccurate because the majority of Zn is cumulative inside cells and is not free in the blood [116]. There are several reasons that Zn is important to men’s health. Assisting immune function, patronage of healthy cell growth, having a role in preserving prostate health, sexual health, and testosterone hormone levels are typical examples. It has been demonstrated that Zn plays a significant role in reproductive functions. Zinc is an indispensable inorganic element universally used in medicine, biology, and industry. Its daily intake in an adult is 8–15 mg/day, of which approximately 5–6 mg/day is lost through urine and sweat. Also, it is an

Metallodrugs in Medicinal Inorganic Chemistry  99 essential constituent of bones, teeth, enzymes, and many functional proteins. Zinc metal is an essential trace element for man, animal, plant, and bacterial growth while zinc oxide nanoparticles are toxic to many fungi, viruses, and bacteria. People with inherent genetic deficiency of soluble zinc-binding protein suffer from acrodermatitis enteropathica, a genetic disease indicated by python like rough and scaly skin. Although conflicting reports have been received about nanoparticles due to their inadvertent use and disposal, some metal oxide nanoparticles are useful to men, animals, and plants. The essential nutrients become harmful when they are taken in excess. The mutagenic potential of zinc oxide (ZnO) has not been thoroughly studied in bacteria even though The DNA-damaging potential has been reported. Impact of zinc oxide on biological functions depends on its morphology, particle size, exposure time, concentration, pH, and biocompatibility. They are more effective against microorganisms such as Bacillus subtilis, Bacillus megaterium, Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumonia, Pseudomonas vulgaris, and Candida albicans. Mechanism of action has been ascribed to the activation of ZnO nanoparticles by light, which penetrates the bacterial cell wall via diffusion. It has been confirmed from SEM and TEM images of the bacterial cells that ZnO nanoparticles disintegrate the cell membrane and accumulate in the cytoplasm where they interact with biomolecules causing cell apoptosis leading to cell death (Figure 3.11).

Increases fertility

Maintains Heart Health by Supporting Blood Vessels

Balances Hormones

Helps with Muscle Growth and Repair Zinc as a health Benefits Aids in Nutrient Absorption and Digestion

Supports Liver Health

Fights Diabetes

Increases Immunity and Fights Colds

Acts as Powerful Antioxidant that May Help Fight Cancer

Figure 3.11  Zinc in human health and benefits.

Prevents Diarrhea

100  Advances in Metallodrugs

3.4.7 Zinc in Alloy and Composites Recently, zinc and zinc-based alloys were proposed as new additions to the list of degradable metals and as promising alternatives to magnesium and iron. The following are advantageous characteristics of zinc and its alloys for use in medical applications: • Similar to magnesium and iron, zinc is an essential trace element in the human body. It is a component of more than 300 enzymes and an even greater number of other proteins, highlighting its indispensable role in human health. Optimal nucleic acid and protein metabolism, as well as cell growth, division, and function, require sufficient availability of zinc. • From this perspective, zinc ions released from the implant during the degradation phase could integrate into the normal metabolic activity of the host without producing systemic toxic side effects. • Zinc exhibits high chemical activity, with an electrode potential (−0.762 V) falling between that of magnesium (−2.372 V) and iron (−0.444 V). Pure zinc metal, therefore, exhibits moderate degradation rates (faster than the slowly degrading Fe and its alloys, but slower than the rapidly degrading Mg and its alloys) due to passive layers of moderate stability, formed by corrosion products. • Zinc and zinc-based alloys are easier to cast and process due to their low melting points, low chemical reactivity and good machinability. For instance, unlike Mg-based alloys, the melting of zinc alloys can more conveniently be performed in air.

3.4.8 Zinc Supplementation as a Treatment It is believed to possess wide antioxidant properties and have the ability to protect against accelerated ageing of skin and muscles when using as a treatment against a number of diseases. In case of injuries, zinc is reported to speed up the healing process among the subjects with known zinc deficiency. Zinc supplements were used to treat the major depressive disorder (MDD) effectively among individuals with zinc deficiency showing an association of zinc deficiency and MDD. Men with moderate to higher zinc intake may have a lower risk for prostate cancer, but the opposite may be

Metallodrugs in Medicinal Inorganic Chemistry  101 true at extremely high doses and long-term supplementation [117, 118]. Zinc supplementation is also observed to help in controlling infection and stunted growth in children less than 5 years of age. In combination with vitamin A, zinc supplementation helps to reduce the infection risks and increase linear growth among children. In contrast, displacement of zinc from zinc-binding structures, e.g., finger structures in DNA repair enzymes, may even be a major mechanism for the carcinogenicity of other metals such as cadmium, cobalt, nickel, and arsenic [119].

3.4.8.1 Zinc Deficiency The deficiency of zinc causes a number of problems. Deficiency usually is caused due to insufficient level of zinc in food or as a consequence due to malabsorption caused by other problems like acrodermatitis enteropathica, anaemia, chronic liver or renal disease, sickle cell anaemia, malignancy, and other chronic illnesses. Diverse symptoms occur due to mild zinc deficiency, leading to a number of clinical outcomes which include depressed growth, diarrhoea, delayed sexual maturation, eye and skin lesion, impaired appetite and altered cognition, impaired host defence properties, defects in carbohydrate utilization and reproductive teratogenesis [120, 121]. It is known that mild zinc deficiency reduces immune action of the body. However, excess zinc also causes reduced immune actions, emphasizing the importance of normal levels of zinc for the stability of the immune system. Individuals who are at risk of zinc deficiency are those of elderly and children’s prevalent mostly in developing nations. It has been estimated that nearly two billion peoples in developing world are suffering from zinc deficiency, which contributes the death of around 800,000 children below 5 years due to persistent suffering of infections and diarrhoea in the world every year. The prevention of disease and reduction in mortality can considerably be reduced by zinc supplements. However, care should be taken for using zinc supplements and check zinc levels of time to time, as its increased levels also interfere with the absorption of other essential micronutrients. Deficiency of zinc is also common among crop plants, mainly in plants growing in high pH soils making them more susceptible to disease [122, 123]. Naturally, zinc is added to soil through the process of rock weathering. However, zinc also adds up to soil by a number of human activities like fossil fuel combustion, mine waste, phosphate fertilizers, limestone manure, sewage sludge, and particles from galvanized surfaces. High zinc levels in soils are also toxic to plants, although toxic cases of zinc are far less widespread. Zinc deficiency causes growth retardation in children stillbirth, spontaneous abortion in pregnant woman

102  Advances in Metallodrugs immunity both mother and body, repeated respiratory infections, alopecia, depression lack of concentration, skin infections, diarrhea, and oral and genital ulcers may occur. Zn acts as a toxic effect against heavy metals and cigarette inflammatory agents. Trace elements play an important role in the male reproductive process because of their high activity at the molecular level, although they are known to exist in the body at very low levels [124]. Zinc as a hormone balancer helps hormones such as testosterone, prostate, and sexual health and functions as an antibacterial agent in men’s urea system.

3.4.8.2 Zinc Toxicity Zinc is an essential element and its requirement is widely known. However, increased zinc absorption causes malabsorption of iron and copper. Free ions of zinc would lead to increased toxicity as solutions containing free zinc ions are highly toxic to plants, invertebrates, and even vertebrate fish. Studies show that solutions which consist even micromolar amounts of free zinc ions are highly toxic. One study shows that six micromolar zinc solution killing 93% of all daphnia in water. The metallic zinc dissolves readily with the hydrochloric acid of stomach and forms corrosive zinc chloride. In one study, one elderly man taking zinc supplement of 8 mg daily were hospitalized for a number of complications like urinary tract problems more often than those taking a placebo. In addition to affecting copper absorption excessive zinc levels of 100–300 mg/day also interfere with cholesterol. Zinc metal fumes after inhaling is known widely to cause metal fume fever (MFF), which is primarily caused due to inhalation of zinc oxide. This is usually an industrial disease, which is affecting the individuals involved in zinc related occupational works. Inhalation of industrial metal fumes which contains zinc particles of size 30 h) and controlled (20–100 pmoles s−1) NO

ON

NH4

Fe

Fe

Na2

Fe

NO

S

ON

S Roussin's red salt (RRS)

S ON

S

NO Fe

Fe

NO

S

NO

Fe

ON ON

NO

Roussin's black salt (RBS)

Figure 6.9  Structure of RRS and RBS.

172  Advances in Metallodrugs

O

C

H3C

NO CH

N

N

C

Ru N

H3C

3

O

O

O

C

N

NO

Ru

N

N

H3CO

CH3

N C

O

O

C

N

3

Ru

N

N

N

N

N O

[(Me2bpb)Ru(NO)(Resf)]

O

O

O

N

N C

O

O

O

NO OCH

O

O [(Me2bQb)Ru(NO)(Resf)]

[(Me2bQb)Ru(NO)(Resf)]

Me2bpb =1,2-bis(pyridine-2-carboxamido)dimethybenzene Me2bQb =1,2-bis(quinoline-2-carboxamido)dimethylbenzene Me2bQb =1,2-bis(quinoline-2-carboxamido)dimethoybenzene Resf = Resorufin dye

Figure 6.10  Dye-attached Ru nitrosyls with high capacities of NO release under illumination with visible light.

durations after visible light irradiation. Quantitation of the efficiency of NO release from the composites shows that polymer gas permeability most dramatically affects the overall efficiency (QY) of photochemical NO release, where polymers with higher gas permeability have a higher QY of nitric oxide release. A prototype LED device shows proof-of-concept that such photoresponsive NO-releasing composites could be applied to implantable systems, where the amount of NO released is modulated by changing irradiation time and light intensity. This research provides the guidelines necessary to move towards device fabrication and testing in actual tissue to evaluate the photo-NORMS as a reliable option for nitric oxide release in vivo. Our research group [145] has shown the photodynamic behavior of ruthenium(II) nitrosyl complex containing N,N’-salicyldehydeethylenediimine (SalenH2) through density functional theoretical (DFT) studies using Gaussian 09W software package. The DFT calculations with B3LYP/LANL2DZ specified for Ru atom and Becke-3–Lee–Yang–Parr (B3LYP)/6-31G(d,p) combination for all other atoms were used using effective core potential method. Both, the ground and excited states of the complex were evolved. In vitro anticancer aspects against COLO-205 human cancer cells have also been carried out with regard to the complex.

NO-, CO-, and H2S-Based Metallopharmaceuticals   173

Ultraviolet

100

200 315

Visible

Infrared

400

700

Etotal = –41.0407 eV

Wavelength(nm)

hν Cell death

Oxygen

Triplet static

Lig

I ISC C

NO

N

ht Ru

O

Ru

N

O

ROS formation or Fession reaction

Etotal = –41.0397 eV ISC

N O

Etotal = –4.7724 eV Etotal = –39.0771 eV

Figure 6.11  Energy-wise depiction of NO release from reactant to product phase.

More emphasis has been laid to extrapolate DFT to depict the chemical power of the target compound to release nitric oxide (Figure 6.11). These results strongly attest the fact that inorganic chemists could play important roles in this area of drug development.

6.6 Exogenous CO Donating Molecules Because of the enormous therapeutic potential of CO, large efforts have been initiated in the past years to develop new ways of delivering this gas to specific tissues and organs. Pharmaceutical CO administration by inhalation requires CO doses that are sufficiently high to allow diffusion of therapeutically effective amounts of CO from blood to tissues. Direct administration of CO by inhalation of the gas is limited by the high affinity of hemoglobin for CO, leading to potentially rapid intoxication owing to the facile formation of high levels of carboxy-hemoglobin. In addition, the direct use of CO gas has several disadvantages: special equipment is required (ventilators, masks, inhalers, etc.) and establishment of

174  Advances in Metallodrugs appropriate doses is a complex process, thus, limiting its utilization to specialized hospital settings. The administration of CO by inhalation can be considered more suitable for treatment of respiratory tract and lung disease or for the treatment of a transplant donor and/or ex vivo treatment of an organ to be transplanted. To surpass the above limitations for effective systemic CO delivery, alternative pharmaceutical approaches have been designed that encompass: (i) the use of pro-drugs that are metabolically converted into CO, and (ii) the use of CO releasing molecules (CORMs), which are able to bind, stabilize, release, and deliver CO. However, in many cases, the administration of CORMs can also cause an increase in carboxy-hemoglobin level, and efforts are needed to develop new molecules that are able to release CO in a tissue-specific manner. The use of pro-drugs (Figure 6.12), such as methylene dichloride (dichloromethane) [146] and organic aldehydes [147] find their use in the selective formation of CO in tissues that possess a specific enzymatic activity, which converts the compound with concomitant release of CO. Provided that a suitable combination of pro-drug and metabolic activity is used, this approach is potentially useful for the tissue-selective targeting of CO release. Thus far, cytochrome P450- and glutathione-S-transferase-­ mediated decarbonylation reactions have been implicated in the release of CO from methylene chloride. The 2nd strategy, based on CORMs, is considered as more promising for therapeutic applications. Three main classes of CORMs with potential pharmacological actions have been described and these are transition metal carbonyls, boranocarbonates, and aldehydes. At the end of the 1990s, CO had emerged as a potential therapeutic molecule, it became necessary to discover solutions to deliver and store CO in a controlled manner, as well as to target it to diseased tissues. This needed to be achieved without interfering with hemoglobin-mediated oxygen transport in the systemic circulation. Developing ways of providing CO in a chemically stable form was imperative. This inspired several groups to identify exogenous

Cl

H Cl

C

R1

C

C R3

O

O

O

H

H

H3C

H

C

H3C R2

C

H3C

C

CH3

Figure 6.12  Methylene chloride and organic aldehydes as CORMs.

C CH3

H

NO-, CO-, and H2S-Based Metallopharmaceuticals   175 CO-donors (CORMs) for controlled delivery of CO to safeguard against cardiac tissue damage, organ graft rejection, and post-operative complications [148–150]. Transition metal carbonyls were found to be promising compounds for releasing CO. These complexes are constituted by a transition metal (such as manganese, cobalt, or iron) surrounded by a certain number of carbonyl groups as coordinated ligands. CORM-1, [Mn2(CO)10] (Figure 6.13), was the first compound to present biological activity related to CO [151, 152]. CO was released from CORM-1 after photo-excitation, which was measured by conversion of deoxymyoglobin into carbonmonoxymyoglobin (MbCO). Additionally, in isolated heart, CORM-1 produced attenuation of coronary vasoconstriction induced by an inhibitor of NO-synthase [153]. Following these first encouraging results, other CORMs have been developed. CORM-2 and CORM-3 (Figure 6.13) are ruthenium-based compounds with formulae [Ru(CO)3Cl2]2 and [Ru(CO)3Cl(glycinate)], respectively [151, 152]. CORM-2 has been shown to induce vasodilatation in isolated rat aorta. CORM-3 is water-soluble (unlike CORM-2) and has become the most studied CORM with many described actions, e.g., it can induce vasodilatation, has anti-inflammatory properties in macrophages and neuronal cells, and protects the kidney from cold ischemia/reperfusion injury. Iron carbonyl, namely, [Fe(CO)2(S-N)2] (CORM-S1, Figure 6.13) [154], has also been reported to exhibit lightdependent CO liberation. The rates of CO release from the rhenium carbonyl [Re(L)2(CO)2Br2] (L = py, im, and other N-donor ligand, Figure 6.13) [154] can be varied via change of the N-donor ligand.

OC

CO

O C

Cl

CO

CO

OC

Mn

Mn OC

CO

C O

Cl

OC

Cl

Cl

Cl

Ru Cl

OC

[Ru(CO)3Cl2DMSO)]

N

Figure 6.13  Examples of CORMS.

N Ru

OC

O

Cl CORM-3

CO

S

Fe

Br

OC

H2 N

Ru CO

OC Cl

CO

OC

CORM-2: [Ru(CO)3Cl2]2

DMSO

OC

CO

Ru

Ru CO

CORM-1: Mn2(CO)10

OC

CO

Cl

Br

OC [Re(py)2CO)2Br2]

S

NH2

N H

CORM-S1

O

176  Advances in Metallodrugs 2– H 2Na+

H

O

B H

O

Figure 6.14  CORM-A1: Disodium boranocarbonate.

CORMs are not restricted to transition metal carbonyls. CORM-A1 (Na2H3BCO2) (Figure 6.14), unlike CORM-3, does not contain a transition metal but is a boronate compound, which releases CO and boric acid when it comes in contact with water [155]. Under physiological conditions, it releases CO at a much slower rate than many transition metal CO complexes. In the presence of myoglobin, CORM-3 transfers CO forming MbCO with a very fast rate (t1/2 = ~ 1 min), whereas CORM-A1 releases CO much slower (t1/2 = ~ 21 min) at pH 7.4. Therefore, it is thought that CORM-3 would be more applicable for therapeutic applications in which CO acts as a prompt signaling mediator, whereas CORM-A1 would be more suitable for chronic diseases, mimicking better a sustained HO-mediated CO formation. This water-soluble CORM induces vasorelaxation in isolated tissues and causes drop in arterial pressure in vivo via slow release of CO. Several recent reviews are available that address these topics [156–159]. New compounds are being developed that deliver CO in a more controlled manner with respect to tissue and pathology-specificity, minimization of carboxy-hemoglobin formation, and velocity of CO release.

6.7 H2S Donating Compounds With the myriad of purported biological actions of H2S, there is growing interest in more precise delivery of this volatile gas to target tissues in the form of H2S donating compounds. H2S donating compounds can be divided into four general classes: H2S gas, sulfide salts, synthetic moieties, and naturally occurring compounds (mainly in foods).

6.7.1 H2S Gas: A Fast Delivering Compound The simplest and most straightforward H2S preparation is H2S itself either in an aqueous form or a gaseous form. The bubbling of a physiological saline with pure H2S gas yields a H2S solution with known concentrations of H2S [160]. The conventional preparation involves bubbling the saline

NO-, CO-, and H2S-Based Metallopharmaceuticals   177 with pure H2S gas at 30°C for 40 min to obtain a H2S-saturated solution (0.09 M) as step 1. Step 2 is to dilute this saturated solution further to the desired final concentration. Treatment of animals, tissues, or cells with this H2S gas-bubbled solution has the advantage for the ready availability of pure H2S gas and the bubbling apparatus as well as the simplicity for interpretation due to the lack of other nonsulfide elements in the solution. As the H2S is dissolved in the liquid for this preparation, the question is often asked as per the release of H2S from the solution. This is a legitimate concern. H2S can be released from the solution in a temperature- and ­concentration-dependent manner. At 37°C or below, the concentration of H2S of the bath solution is found to be stable. When the H2S-bubbled solution has the initial H2S concentration below 1 mM, 15% decrease in H2S concentration in the solution would occur within 30 min [160]. Therefore, in most controlled experiments in a laboratory setting with room temperature around 22 or 37°C for cell culture experiments, the H2S-bubbled solution can be reliably used as long as the solution will be refreshed frequently. One obvious drawback for the bubbled solution is the requirement of a well-ventilated environment that will quickly remove the discharged H2S gas from the bubbling process, which sometimes becomes problematic.

6.7.2 Sulfide Salts: Fast Delivering H2S Compounds Sodium hydrosulfide (NaHS) has been used as a H2S donor more often than any other donors including H2S-bubbled solution. The NaHS standard solution can be made by logarithmically diluting from freshly made 1 M NaHS stock solution. There are several reasons behind its wide utilization. One is its convenient preparation without pungent smell during the preparation process, simply adding NaHS compound to the liquid. Another one is based on the dissociation of NaHS into Na+ and HS− in solution. The latter can associate with H+ to form H2S. Given a physiological pH around 7.4 and temperature of 37°C, NaHS solution will yield about one-third of the undissociated H2S and the other two-third remains as HS−. Therefore, it is conventionally believed that the NaHS solution will give the researchers better control of the concentration of H2S actually delivered than H2Sbubbled solution. However, one has to be very careful that the dissociation of NaHS is pH dependent and the generated H2S can also be released from the solution in a temperature- and concentration-dependent manner. Sodium sulfide (Na2S), under the name of IK-1001, has been used as a H2S donor in a number of studies [161–163]. This salt is soluble in water, but not in ethanol as sodium hydrosulfide (NaHS) is. This differential solubility can be used to separate NaHS from Na2S and sodium thiosulfate.

178  Advances in Metallodrugs S

S

O

S

P

P

S

O

NH2 HO

S (a)

(b)

Figure 6.15  (a) Lawesson’s reagent and (b) 4-hydroxythiobenzamide.

Once in solution, Na2S dissociates and generates H2S. In addition to having one more choice in using H2S donors, it is not clear whether Na2S is superior to other donors [164]. It has not been determined how much H2S would be released at a pH of ~7.4. Lawesson’s reagent [2,4-bis(4-methoxyphenyl)-1,3,2,4-dithiadiphosphetane 2,4-disulfide], (Figure 6.15a) originally used in organic synthesis as a thiation agent [164], has been applied in biological studies as a H2S donor in few reports. This reagent (0.1–3 µmol/kg) has been used to assess the effects of H2S on aspirin- and fMLP-induced leukocyte adherence [163] (755), on neutrophil migration [164], on carrageenan-induced knee joint synovitis [165], and on ulcer healing [166]. Another H2S donor reported is 4-hydroxythiobenzamide (Figure 6.15b) (4-HTB) [167, 168]. Due to the limited studies, it is difficult to assess its advantages or disadvantages over other H2S donors.

6.7.3 Synthetic Moieties 6.7.3.1 Slow-Delivering H2S Compounds The quick delivery of H2S by H2S-bubbled gas or NaHS challenges the situations where a long-term sustained supply of H2S is required and also where a fast H2S overshoot surge followed by rapid decline is undesirable. To answer this situation, H2S-releasing compounds have been developed in the last several years, and some of these slow-releasing compounds are GYY4137 [169] and S-diclofenac [170, 171]. Compared to Lawesson’s reagent and sulfide salts, GYY4137 [­morpholin4-ium 4-methoxyphenyl-(morpholino)-phosphinodithioate] can generate H2S at a lower rate. It releases H2S slowly and steadily, either in aqueous solution or administered to the animal (intraperitoneal or intravenous) [169]. The slow-releasing property of GYY4137 results in slow vasorelaxation without affecting heart rate or myocardial contraction of rats.

NO-, CO-, and H2S-Based Metallopharmaceuticals   179 S O

P

S– HN+

O

N

O

Figure 6.16  Structure of GYY4137.

Also, due to this slow-releasing profile, GYY4137 (Figure 6.16) inhibited the development of hypertension in a NG-nitro-L-arginine methyl ester (L-NAME)-evoked rat hypertension model or in SHR rats, after 14 days of administration [169]. While NaHS is pro-angiogenetic [172], H2S slow-releasing S-diclofenac inhibits angiogenesis and endothelial cell proliferation [173].

6.7.3.2 H2S-Releasing Composite Compounds H2S-releasing composite compounds are being developed by combining a H2S-releasing moiety with another parent compound with known molecular structure and biological functions. The purpose behind developing such a new compound is to augment the functionality and safety of both compositing compounds and reduce potential side effects of each moiety. To date, this purpose appears to be partially achieved from animal studies. The challenges for these H2S hybrids molecules include the analysis of the pro and con for their chronic usage, H2S releasing kinetics, metabolic dynamics in vivo, and accessibility to different organs and tissues. In fact, the final test of their usefulness would be the clinical trials. Principally, the H2S-releasing composites are made by grafting onto existing compounds, the moiety 5-(4-hydroxyphenyl)- 3H-1,2-dithiole-3-thione (ADTOH), the main metabolite of ADT [174]. Some examples of these newly made-up H2S moieties are reviewed below.

6.7.3.2.1 S-Aspirin: ACS14

It is a composite compound of aspirin and ADTOH. Aspirin has the tendency to cause gastric damage while protecting the cardiovascular system. ACS14 is found to inhibit thromboxane synthesis, similar to aspirin, but caused much weaker gastric reactions than aspirin [175]. This could be due to the antioxidant behavior of H2S released from the composite compound. Another compound with S-aspirin is ACS21 (S-salicylic acid). Both ACS14

180  Advances in Metallodrugs O O S O S

S

O

Figure 6.17  Structure of ACS-14.

and ACS21, but not aspirin and salicylic acid, limited the development of metabolic syndrome in rats induced by GSH depletion (including hypertension, endothelial dysfunction, and insulin resistance), and protected the heart from I/R damage. Moreover, no gastric lesion was found with the application of ACS14 or ACS21 [176, 177]. Oral administration of ACS14 (Figure 6.17) for 7 days or a single intraperitoneal injection to rats significantly increased GSH levels in heart, aorta, and plasma [178, 179]. This outcome on thiol metabolism was also achieved with ADTOH, but was not with aspirin.

6.7.3.2.2 S-Sildenafil: ACS6

ACS6 (Figure 6.18) is a hybrid compound of sidenafil and ADTOH. Sildenafil is an efficient and selective inhibitor of type 5 phosphodiesterase (PDE5), of which the location in corpus cavernosum and prostate is especially related to abnormalities in male reproductive system and urinary O O

CH3 H N O H2C

S

O

O2S

O

N N

H3C

N

H2C H3C

CH2

Figure 6.18  Structure of ACS-6.

CH3

S

S

NO-, CO-, and H2S-Based Metallopharmaceuticals   181 system. With increased oxidative stress level and NOX expression, such as in diabetes, the efficiency of sildenafil in treating erectile dysfunction is lessened. Both ACS6 and sildenafil relaxed cavernosal smooth muscle, but ACS6 is more potent in inhibiting the formation of superoxide and expression of p47 (phox) (a protein composed of 390 amino acids with a molecular mass of 44.7 kDa) and PDE5 than sildenafil [180]. The S-sildenafil may also find its application to those patients with erectile dysfunction, who do not respond to sildenafil treatment. Furthermore, NaHS and ACS6 both inhibited superoxide formation in cultured porcine pulmonary arterial endothelial cells, but ACS6 is about at least 10 times more potent than NaHS [181].

6.7.3.2.3 S-Latanoprost: ACS67

ACS67 (Figure 6.19) is a composite compound of H2S donating moiety dithiolethione (ACS1) and latanoprost. Latanoprost is a synthetic derivative of the natural prostaglandin F2α. As an agent to lower intraocular pressure (IOP) in glaucoma management, latanoprost does not protect retina from ischemia damage, and its tolerance by patients is low. Among retinal damages after ischemia are altered electroretinogram (ERG), reduced retinal localization of specific antigens, and decreased optic nerve axonal proteins. Intravitreal injection of ACS67 immediately after ischemia blunted most of these abnormalities. ACS67, but not of latanoprost, attenuated the death of cultured retinal ganglion cells (RGCs) by increasing GSH levels and decreasing H2O2 toxicity [182]. This observation from cultured cells was supported by the whole animal study in glaucomatous pigmented rabbits. In these animals, ACS67 achieved a greater anti-IOP effect than latanoprost. Moreover, the rabbits tolerated ACS67 well over a 5-day treatment regimen with daily intraocular administration [183]. The argument is made that the additional neuronal protective effect of ACS67 compared with latanoprost is due to the release of H2S from the hybrid. HO

O O S

HO OH

Figure 6.19  Structure of S-Latanoprost: ACS67.

S

S

182  Advances in Metallodrugs

6.7.3.2.4 HS-NSAIDs Hybrid Compounds

The use of nonsteroid anti-inflammatory drugs (NSAIDs) suffers from unacceptable risk of gastrointestinal ulceration and bleeding [184–187]. In order to reduce such side effects, 1,2-Dithiole-3-thiones (DTTs) have been conjugated to NSAIDs to form HS-hybrid NSAIDs (HS- NSAIDs), which showed significant reduction of gastrointestinal damage compared to the parent NSAIDs [188, 189]. In addition, HS-NSAIDs also boosted the anti-­ inflammatory effect of their NSAIDs counterparts. In a work by Fiorucci et  al. [190], DTT was conjugated to diclofenac to afford a HS-NSAID-hybrid ATB-337, and its anti-inflammatory effect was investigated along with diclofenac in rats. In a rat air pouch model, orally administrated ATB-337 dose-dependently suppressed the activity of both COX-1 and COX-2, and the efficiency was comparable to that of the diclofenac. Additionally, pretreatment with ATB-337 and diclofenac led to a reduction of ­carrageenan-induced paw swelling volume. It is notable here that pretreatment with ATB-337 at 10 µmol/ kg achieved a reduction in edema formation comparable to that seen with diclofenac at 30 µmol/kg. This enhanced potency was probably associated with the generation of H2S from ATB-337. An enhanced anti-inflammatory effect was also observed for ATB-429 [191]. In addition to their anti-inflammatory effect, other HS-NSAIDs including HS-sulindac (HS-SUL), HA-aspirin (HS- ASA), HS-ibuprofen (HS-IBU), and HS-naproxen (HS-NAP) were also reported to exhibit anti-proliferative effect against human colon, breast, pancreatic, prostate, lung, and leukemia cancer cell lines. The conjugation with 5-(4-­ hydroxyphenyl)-1,2-dithiol-3-thione (ADT-OH) significantly increased the growth inhibitory effect of NSAID by 28- to > 3000-fold [192].

6.7.4 Naturally Occurring Plant Derived Compounds In our daily life, many dietary elements directly and indirectly provide H2S to our body so that the advantage of this gasotransmitter can be realized, in many cases, without conscious thought. Some of these natural nutritional sources for H2S supplementation are given here.

6.7.4.1 Garlic Garlic (Allium sativum) assists in lowering blood pressure and protecting the heart [193], fighting with atherosclerosis, lowering blood sugar and cholesterol levels [194], and preventing platelet aggregation. Garlic has also been found to be effective against bacterial, viral, fungal, and parasitic

NO-, CO-, and H2S-Based Metallopharmaceuticals   183 infections, in addition to enhancing the immune system and having antitumoral and antioxidant features [195]. Role of garlic in the prevention of ischemia-reperfusion injury [196], as anticancer activity for breast cancer [195] and in improvement of insulin sensitivity and associated metabolic syndromes in rats [196] have also been reported. For a long time, the pungent aroma of garlic has been noticed and attributed to volatile sulfur-containing flavor compounds, but how much credit of the health benefits of garlic should be given to these volatile compounds and what are their final sulfur products have not been clear. While garlic has long been felt beneficial as an antioxidant, recent evidence suggests that a number of beneficial effects of garlic are derived from H2S [197] production. Thus far, the best characterized naturally occurring H2S-donating compound from garlic (Allium sativum) is allicin (Figure  6.20) (diallyl thiosulfinate) which decomposes in water to a number of compounds [198]. Benavides et al. [199] measured H2S production in real time with a polarographic sensor and observed that under anoxic conditions and in the presence of GSH, red blood cells rapidly (within minutes) produced H2S from garlic extract, as well as from two of the decomposition products of allicin, diallyl disulfide (DADS) and diallyl trisulfide (DATS) (Figure 6.20), with approximately 3 times the yield from the latter. H2S could also be generated in the presence of GSH, without red blood cells, and by reduced thiols on the red blood cell membrane. Infusion of diallyl disulfide 1.8 mg/kg/min in rats increased exhaled H2S providing additional support for H2S production from garlic in vivo. Other molecules that can be extracted from garlic include a number of analogs of cysteine that are also readily synthesized. Ajoenes are also potential H2S donors, although H2S production from these molecules has not yet been measured. O S

S

Allicin

S

S S Diallyl disulfide (DADS)

Figure 6.20  Structure of Allicin, DADS, and DATS.

S S

Diallyl trisulfide (DATS)

184  Advances in Metallodrugs

6.7.4.2 Broccoli and Other Cruciferous Vegetables Sulforaphane (Figure 6.21), the isothiocyanate compound from broccoli (Brassica oleracea), is rapidly absorbed by humans, reaching peak concentrations at 1 h and declining thereafter with a half-life of 1.8 h [200, 201]. A related isothiocyanate compound, erucin (Figure 6.17), is found in high levels in rocket salad species (Eruca sativa). Allyl isothiocyanate (Figure  6.17) is derived from wasabi, mustard, and horseradish. It has not yet been determined if any of the isothiocyanates are metabolized to produce H2S, although their purported health benefits are quite similar to those that are attributed to H2S or other H2S donating drugs, suggesting a role for H2S, although this needs to be evaluated further (Figure 6.21). Broccoli as well as other cruciferous vegetables, such as cabbages, turnips, and wallflowers, tends to release strong smell of H2S when cooked or became rotten. Previous studies have reported the health beneficial effects of broccoli on the prevention and treatment of hypertension and atherosclerotic changes in the SHR stroke-prone rats [202]. These effects of broccoli are largely credited to the action of sulforaphane that exhibits anticancer properties, and young sprouts of broccoli are particularly rich in sulforaphane. In a recent study by Pei et al. [203], the release of H2S from sulforaphane was unmasked under different experimental conditions, such as the addition of sulforaphane to the cell culture medium or mouse liver homogenates or administration of sulforaphane to the living mice. While sulforaphane speeded the death of PC-3 cells (a human prostate cancer cell line) in a dose-dependent manner, scavenging of free H2S with methemoglobin or oxidized glutathione reversed sulforaphane-reduced cell viability. The anti-prostate cancer effect of sulforaphane was also mimicked by NaHS treatment as PC-3 cell migration was inhibited by NaHS. The activation of p38 MAPK and c-Jun NH2-terminal kinase (JNK) appears to be the common signaling linkage for the effect of sulforaphane and NaHS N C

S O

S

N C S

Erucin

sulforaphane N C

S

Allyl isothiocyanate

Figure 6.21  Structure of some isothiocyanate compound.

S

NO-, CO-, and H2S-Based Metallopharmaceuticals   185 in PC-3 cells. Suppression of both p38 MAPK and JNK reversed NaHS- or ­sulforaphane-reduced viability of PC-3 cells. Sulforaphane, Erucin, and a related isothiocyanate, iberin, pregulate thioredoxin reductase 1 (EC 1.8.1.9) in human breast cancer MCF-7 cells. Because thioredoxin reductase is the only enzyme that reduces thioredoxin and reduced thioredoxin is required to release H2S from 3-MST, it is quite possible that many of the health benefits of cruciferous vegetables involve H2S. Sulforaphane protects vascular SMCs and endothelial cells from oxidative and inflammatory stress and suppresses angiogeniesis [204–206]. Sulphoraphane has neuroprotective and anti-inflammatory actions mediated in part through activation of heme oxygenase-1 (HO-1) and it provides some protection against ischemia reperfusion injury, hemorrhage and serotonin induced toxicity [207, 208].

6.8 Concluding Remarks and Future Outlook Both CO and NO are essential signalling molecules in the body. H2S is now labeled as the 3rd signalling molecule after NO and CO. The importance of NO was discovered in the 1970s and has led to a much greater understanding of the biochemistry of NO and the development of NObased pharmaceuticals. To date, the use of NO in medicine has been limited to slow release of low doses into the blood stream through decomposition of various exogenous systemic NO donors. However, site-specific delivery of this reactive gas in high doses to combat chronic infection and neoplasms still remains a challenge. On account of the later discovery that CO is important, the understanding of the role of CO is lagging more than two decades behind NO. Most of the researches on CO have been concentrated in the areas of the biological and medical effects of CO. However, both designing novel CO-based drugs and understanding of CO and CORM biology are interdependent tasks that must progress side-by-side. This will provide an exciting scientific area for the collaboration between inorganic chemists, biologists, pharmacologists, medicinal chemists, and physicians who may highlight the unmet medical needs where CO therapy may provide a desirable breakthrough. The range of diseases that are responsive to CO together with the ongoing elucidation of the methodology required to deliver CO to specific disease sites is paving the way for the use of CO-based drugs in the clinic. It is probable that CO will be used in human medicine in the near future with an important role for CO-releasing molecules.

186  Advances in Metallodrugs In the recent past, there has been growing research interest in H2S as a signalling molecule. As H2S has a pKa value of 6.8, it is mainly present as H2S and [HS]− at physiological pH. It is more toxic than CO. In the brain, H2S is synthesized from cysteine, catalyzed by cystathionine β-synthase (CBS) cystathionine-γ-lyase (CSE), and 3-mercaptopyruvate sulfurtransferase (3-MST), but it is probable that there are other pathways. H2S concentration is 50–160 mM in brain tissue and 10–100 mM in blood, which is much higher than is found for either CO or NO. H2S is active in both the cardiovascular and central nervous systems. As [HS]− readily coordinates to hemes and some cytochromes, it is not surprising that it is biologically active, but more research is needed to establish whether it is an important signalling molecule or not. Many studies have proven the clinical relevance of the molecules in question, but the arena of human clinical trials is yet to be opened. Besides treating dreadful diseases like cancer and diabetes by their virtue, a major void is yet to be filled to create a flawless disease controlling strategies. Our research groups are also searching for the same class of metallopharmaceuticals to meet the suitable molecular requirements of medicinal purview [209–216].

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NO-, CO-, and H2S-Based Metallopharmaceuticals   199 185. Singh, G., Fort, J.G., Goldstein, J.L., Levy, R.A., Hanrahan, P.S., Bello, A.E., Celecoxib versus naproxen and diclofenac in osteoarthritis patients: SUCCESS-I study. Am. J. Med., 119, 255–266, 2006. 186. Kurahara, K., Matsumoto, T., Iida, M., Honda, K., Yao, T., Fujishima, M., Clinical and endoscopic features of nonsteroidal anti-inflammatory drug-­ induced colonic ulcerations. Am. J. Gastroenterol., 96, 473–480, 2001. 187. Sparatore, A., Santus, G., Giustarini, D., Rossi, R., Del Soldato, P., Therapeutic potential of new hydrogensulfide-releasing hybrids. Expert Rev. Clin. Pharmacol., 4, 109–121, 2011. 188. Chan, M.V. and Wallace, J.L., Hydrogensulfide-based therapeutics and gastrointestinal diseases: Translating physiology to treatments. Am. J. Physiol. Gastrointest. Liver Physiol., 305, G467–473, 2013. 189. Wallace, J.L., Caliendo, G., Santagada, V., Cirino, G., Fiorucci, S., Gastro­ intestinal, safety and anti-inflammatory effects of a hydrogensulfide-releasing diclofenac derivative in the rat. Gastroenterology, 132, 261–271, 2007. 190. Fiorucci, S., Orlandi, S., Mencarelli, A., Caliendo, G., Santagada, V., Distrutti, E., Santucci, L., Cirino, G., Wallace, J.L., Enhanced activity of a hydrogen­ sulphide-releasing derivative of mesalamine (ATB-429) in a mouse model of colitis. Br. J. Pharmacol., 150, 996–1002, 2007. 191. Chattopadhyay, M., Kodela, R., Nath, N., Dastagirzada, Y.M., velazquez-­ Martinez, C.A., Boring, D., Hydrogensulfide-releasing NSAIDs inhibit the growth of human cancer cells: A general property and evidence of a tissue type-independent effect. Biochem. Pharmacol., 83, 715–722, 2012. 192. Benavides, G.A., Squadrito, G.L., Mills, R.W., Patel, H.D., Isbell, T.S., Patel, R.P., Darley-Usmar, V.M., Doeller, J., Kraus, D.W., Hydrogen sulfide mediates the vasoactivity of garlic. Proc. Natl. Acad. Sci. U.S.A., 104, 17977–17982, 2007. 193. Mukherjee, S., Lekli, I., Goswami, S., Das, D., Freshly crushed garlic is a superior cardioprotective agent than processed garlic. J. Agric. Food. Chem., 57, 7137–7144, 2009. 194. Corzo-Martínez, M., Nieves, C., Villamiel, M., Biological properties of onions and garlic, Trends. Food. Sci. Technol., 18, 609–625, 2007. 195. Sener, G., Sakarcan, A., Yegen, B.C., Role of garlic in the prevention of ischemia-reperfusion injury. Mol. Nutr. Food Res., 51, 1345–1352, 2007, [PubMed: 17966137]. 196. Tsubura, A., Lai, Y.C., Kuwata, M., Uehara, N., Yoshizawa, K., Anticancer effects of garlic and garlic derived compounds for breast cancer control, Anticancer. Agents Med. Chem., 11, 249–53, 2011, [PubMed: 21269259]. 197. Padiya, R., Khatua, T.N., Bagul, P.K., Kuncha, M., Banerjee, S.K., Garlic improves insulin sensitivity and associated metabolic syndromes in fructose fed rats. Nutr. Metab. (Lond.), 8, 53–61, 2011, [PubMed: 21794123]. 198. Kashfi, K. and Olson, K.R., Biology and therapeutic potential of hydrogen sulfide and hydrogen sulfide-releasing chimeras. Biochem. Pharmacol., 85, 689–703, 2013.

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7 Platinum Complexes in Medicine and in the Treatment of Cancer Rakesh Kumar Ameta* and Parth Malik School of Chemical Sciences, Central University of Gujarat, Gandhinagar, Gujarat, India

Abstract

This chapter provides a thematic understanding of platinum (Pt) chemistry with the intent to portray salient biomedical aspects of therapeutically active Pt complexes. Major focus has been laid on understanding the Pt complexes formation with multiple ligands conferring potent anticancer traits alongside reducing its fatality. The accidental discovery of cisplatin (a Pt complex) laid the platform for exploring Pt in medicinal domain, as a result of which, the corresponding chemistry is being explored persistently, since the past several decades. Studies on modulating the design of Pt complexes for bettering their consequent anticancer potency point out a critical role of metal-ligand interactions. The cell specific responses of these complexes have elucidated manifold permutations and combinations of their workable models. Concurrently, a role of structure-activity relationships (SAR) is deciphered as a crucial link to accomplish the need based performance extents. With such enthusiasm, the present chapter attempts to discuss the explored and likelihood functional aspects of Pt complexes in medicinal domain. Keywords:  Cancer, Pt complex, DNA binding, structure-activity relationship

7.1 What is Cancer? Among the most primitive growth indications, formation of new cells from previously existing cellular populations (as per the requirement of different *Corresponding author: [email protected] Shahid-ul-Islam, Athar Adil Hashmi and Salman Ahmad Khan (eds.) Advances in Metallodrugs: Preparation and Applications in Medical Chemistry, (203–246) © 2020 Scrivener Publishing LLC

203

204  Advances in Metallodrugs physiological domains) forms the most essential aspect. In general, cells perish either after a definite life span (cessation in their active functioning) they become too old or upon being irreversibly damaged by external injuries (may be accidental or metabolically driven). The worn out cell populations are compensated by newly formed cells, over a gradual time span, as an essential discourse of homeostatic mechanism [1, 2]. Attenuation in this normal living span of cells within physiological locations, initiates cancer, caused on account of instantaneously activated genetic changes. The hallmark of such physical indications is the uncontrolled cellular growth, characterized by the formation of a solid mass, recognized as tumor. Most common abruptions leading to onset of cancer include either the genetic alterations, that could be hereditary or due to unfavorable working conditions. As an instance, a unanimous threat enriching literature in this aspect is the tobacco smoke finding increasing interest in understanding its chemical diversity propelling it as a cancer causing agent [3, 4]. Tumors depict a normal phenotypic expression of a growth pattern which could be either malignant or benign, the former described by the uncontrolled cellular mass (or population), capable of metastasizing to nearby as well as distinct locations alongside anchorage independent growth and self blood-vessel formation. Benign tumors on the other hand are the ones capable of growing out of proportion but are not metastasized. Thereby, benign tumors are non-metastatic and can be cured (even prevail as normal features of a healthy organism) contrary to malignant counterparts, which possess a rampant metastatic activity alongside aggressively altered phenotypes [5–7]. Of note, all cancers are not known to initiate through tumor formation (such as leukaemias, lymphoma, and myeloma). On the basis of their initiation source (recognizing cells as basic building blocks of life), cancers are normally distinguished into four distinct categories, namely: (a) Carcinomas, (b) Sarcomas, (c) Leukemias, and (d) Lymphomas [8, 9]. Figure 7.1 depicts the characteristic features of these different cancer categories, in terms of their respective initiating organs. In order to understand the unique cancer pathophysiology (for safer and accurate treatment design), it is very important to know about the growth mechanisms and characteristics distinguishing cancer cells from their normal counterparts. In course of a cancer cell growth, the bloodstream transports tumorigenic factors to nearby or distant locations (healthier tissues) other than the originating site. This process aggravates the growth of more and more cancerous cells. Since blood circulation is implicitly regulated by the lymphatic system, so this transport at the very first stage leads to invasion of cancer to lymph nodes (typically, the precursors of white blood cells responsible in making up requisite immune responses) [9, 10].

Platinum Complexes in Medicine  205 Carcinomas: Prevail in skin or tissues forming surface of internal organs and glands, such as breast, lung, prostate and colorectal

Sarcomas: Supporting and connecting tissues of the body such as fat, muscles, nerves, tendons, bone, lymph vessels

Diversity of cancers (on the basis of originating organ)

Leukemias: Cancers of blood, characterized by uncontrolled growth of healthy blood cells, four major types are commonly known

Lymphnmas: Begin from lymphatic system (precursors of immune cells) Hodgkin and nonHodgkin are two main types

Figure 7.1  Different types of cancer, on the basis of respective organs of initiation.

In general, lymph nodes reside as clusters in multiple tissues across the body such as neck, groin area, and under the arms regions. Yet another domain of interest is the understanding of some specific terminologies, such as atypical cells, hyperplasia, dysplasia, and neoplasia [11, 12]. Unlike normal cells, some cells possess characteristic features that are neither cancer nor normal cell like. These cells are atypical cells, having a significant likelihood to become cancerous in near or little distant future. Hyperplasia refers to an abnormal rise in cell population in a tissue or organ. This state weakens the overall immune response, which in turn results in enhancing the vulnerability towards carrying manifold other cancer types. Dysplasia refers to the typical scenario of organ specific enhanced atypical cell population, developed in response to a viral infection (basically a characteristic state between normal and cancer cells). Neoplasia is characterized by the uncontrolled cellular growth, where the involved cells could be benign or malignant.

7.1.1 Characteristic Features of Cancer Cells Two fundamental features of all cancers are unlimited or abnormal cellular growth and the ability to get spread to nearby and distant surroundings (metastasis). The first of these specified responses is stemmed from no restrictive or check-point control on the cell division events compared

206  Advances in Metallodrugs to normal cells (in a same time span) apart form their own blood vessel formation (angiogenesis) for nutrient and serum transport. The specific process of altogether native blood vessel formation in the cancer cells is termed as angiogenesis. Apart from this, cancer cells exhibit significant variations in their cross-communication with extra cellular matrix (ECM) proteins (contrary to normal cells) apart from growing in an anchorage independent manner [13, 14]. In nut shell, the following factors describe the implicit indications of a cell to be considered as cancerous: (i) (ii) (iii) (iv) (v)

Missing regulatory apoptosis mechanism. Continuous growth and division in every circumstances. Invasion of near and distant physiological locations. Anchorage independent growth. Formation of new blood vessels.

7.1.2 Definition of Anticancer Compound In most generalized sense, the growth of any cell (including the cancer) critically depends on the systematic pacification of its DNA replication. Relying on this, if any potential compound is able to cease this replication via interaction, covalent or coordinate bonding (with DNA), it could be considered as anticancer. Holding such features, Pt complexes have emerged as most effective towards curing the cancer, thanks to their co-ordinate bon forming ability to disrupt the native base pairing between constituent base pairs. Contrary to cancer cells, the normal cells perpetuate by growing, dividing and then perishing after a maximum growth. A prominent feature of cancer cells is the continuation of growth and further cell-division instead of normal perishing, further increasing the new abnormal cells. DNA damage is reported as one of the major cancer cause, gradually rendering the inability of native repair mechanism to correct them. Notable aspects among other causative factors could be either environmental or habitual, such as consuming tobacco, diet pattern manifested obesity, acute or chronic exposure to ionizing/non-ionizing radiations, stress, lack of physical activity, and occupational exposure to pollutants [15, 16]. Chemotherapy is the process of treating a disease (most usually cancers) using chemicals having a specific toxicity mediated via regulated or masked expression of microbial activities (antibiotics), owing to which the growth of cancerous tissue is selectively abolished (anticancer therapy). For this procedure, mainly three conventional approaches have been developed, i.e., surgery, radiotherapy, and chemotherapy [15]. First successful neoplasm treatment was developed via surgical removal of tumor

Platinum Complexes in Medicine  207 and its nearby tissues. This procedure was of little assistance pertaining to non-localized tumors (leukemia), unable to arrest the spread (through blood or lymphocytes) to other body locations. Radiation therapy is the curative mechanism facilitating selective elimination of cancer cells. In late 1940s, when nitrogen mustard was used for leukemia treatment, the dawn of chemotherapy descended [17]. Some years later, the field of cancer chemotherapy evolved through more than 30 drug developments, encompassing many diverse chemical moieties, such as Pt coordination complexes, especially, amine ligands-based Pt complexes.

7.1.3 Anticancer Attributes of Pt Complexes Compounds of Pt offer unique possibilities for the design of anticancer chemotherapeutic agents. The manifold possibilities of coordination numbers and geometries, accessible redox states, thermodynamic and kinetic characteristics, and the intrinsic properties of cationic metal ion and ligand itself offer a wide reactivity spectrum that can be exploited through multiple modulating mechanisms. The interest in harvesting anticancer potential of Pt-coordinated complexes arises from their manifested cytotoxic properties. It is pertinent to recall here that being a transition metal, Pt can form complexes with electron donating species, the typical characteristics of extensive natural compounds (imparted by pi-conjugated rich structures) and most of the nitrogen containing derivatives and sub-fractions (like heterocyclic compounds) [18–20]. These complexes are well-documented for disrupting the DNA replication in cancer cells but also work synergistically with manifold chemotherapeutic drugs alongside their increasing referrals as being references standards (pertaining to cytotoxicity induction). Coordinated complexes of Pt with curcumin, paclitaxel, doxorubicin, and plant extract capped metal nanoparticles are already well-known [21–24]. The functional activities of these compounds are in significant focus to improvise the medium dependent biological activities of targeted drugs, such as curcumin [25–27]. Designing such new modalities of antitumor metal-based drugs provides manifold strategies for improving the chemotherapeutic effectiveness of synthetic organic compounds. The underlying working interfaces are defined by molecular mechanisms associated with biological effects of the existing agents. The first and most important task in understanding the functional basis of antitumor agents is the determination of their biological target(s) alongside their characteristic response towards structural recognition by other cellular components. However, anticancer effects are characterized by complex processes (cellular uptake, pharmacokinetics of administered drugs, toxicity, and resistance), each of

208  Advances in Metallodrugs which is further modulated through distinct targets and intervening cellto-cell cross talks. Ligands of such compounds mandate a careful selection for expression of desired toxicity and minimization of undesired side effects. Thus, for understanding the underlying anticancer mechanisms, it is of paramount importance to identify their cellular (pharmacological) targets in compliance with the peculiar chemical environment dependent cytotoxicity. The target moiety (for further discussion) in this compilation has been considered as the characteristically overexpressed receptor (protein) on the cancer cell surface (extensively, a bio macromolecule), which upon interaction with drug facilitates its internalization inside the tumor cells. It is essential to clarify this because a number of studies confuse the intended meaning of target molecule, some terming it as serum protein while others presuming it as a randomly expressed cellular recognition protein/peptide molecule. Nevertheless, the mere fact that such responses alter the native physiological events has often been understood casually in context with possible cytotoxic effects. The observation that the administered anticancer agent modifies properties of the targeted bio macromolecule (through its manifested cytotoxicity), no longer necessitates the cytotoxicity understanding with respect to pharmacological target. This is so as only selective modifications culminating in cell death are included with respect to working actions of potential anticancer agents. Thus, some potential drug candidates in examination might encounter difficulties in the clinical trials just because of inaccurate target identification, in course of their action. Present compilation attempts to address such misconceptions, with high tumor cell toxicity of Pt complexes alongside their interactions with principal cellular target. Hence, threshold toxicity induction restricted to specific sites is fundamental for anticancer activity of as formed complexes (propagated via sustained and chemical environment specific bio-molecular interactions). For instance, targeting of a peculiar cell receptor on different cancers at different stages of their manifestation does not necessarily yield similar and identical intermediary species.

7.1.4 Native State Behavior of Pt Complexes Pt(II) coordination compounds comprised a major proportion of earliest anticancer efficacy optimization work on transition metal complexes. For the very first time, the square-planar geometry of these entities was determined through the standardized isomer-counting experiments, demonstrated by Werner [28]. Subsequently, Pt(I1) complexes were ubiquitously used to elucidate the mechanisms of square-planar substitution reactions,

Platinum Complexes in Medicine  209 Cl

NH3 Pt

Cl

NH3

Figure 7.2  Structure of cisplatin: first anticancer Pt drug.

owing to their reactions at feasible rates in aqueous solution and relatively stable +2 oxidation state confirmations [29]. The binding of Pt to nucleic acids (NA) was first investigated in part with the aim of developing heavy metal stains for electron microscopic (EM) studies on DNA. A high electron density of Pt complexes has also fueled their use as isomorphic replacements for macromolecular crystal structure analysis. Examples of these applications include DNA visualization using Pt shadowing in EM studies [30]. The use of Pt(II) complexes in determining characteristic NA crystal structures has been practiced since ages, to decipher several interfaces, namely, tRNA [31], the nucleosome core particle [32], and the first restriction enzymeDNA substrate complex [33]. Discovery of cisplatin antitumor activity encouraged the exploration of aqueous state Pt coordination chemistry alongside the presumptive interactions (Figure 7.2). Subsequent investigations led to several major advances in basic Pt chemistry, including characterization of previously unknown aqueous Pt and –OH state complexes [34–36], Pt blues and similar polymeric mixed-valence Pt complexes [37–43], Pt-ascorbate complexes [44, 45], and many novel Pt-nucleobase complexes [46]. Gradual interest in this field also bettered the understanding of Pt chemistry in + 3 oxidation state [47] as well as in combination with biologically relevant sulfur ligands [48–51]. Nevertheless, covering details of all such advances is indeed infeasible in the present compilation. Furthermore, these increasing multidisciplinary applications approximate the extent to predict the fundamental inorganic coordination chemistry, when a biological application sparks interest with the concurrent transition metal involvement.

7.2 Compatibility of Pt Compounds in Cancer Treatment 7.2.1 Significance of DNA as Primary Target Coordination chemists have long been fascinated with cisplatin anticancer potency, presumably with a better response of cis than trans configuration.

210  Advances in Metallodrugs Such distinction calls for a stereochemistry conferred preferential interaction of cis isomer with the biological targets, not shared by trans isomer. The DNA remains a well-recognized biomolecule as cisplatin target, overcoming the challenges in understanding the stereochemistry based differences of cis- and transplatin DNA adducts. First evidence of cisplatin biological activity at the DNA level emerged from studies demonstrating its persistent inhibition of DNA synthesis compared to its RNA or protein synthesis impacts [52]. Further proof for DNA level cisplatin activity was inferred from the cisplatin DNA repair sensitivity devoid prokaryotic and eukaryotic cells than their parental cell-lines [53–57]. These experiments inferred a reduction in drug cytotoxicity subsequent to the removal of cisplatin-DNA adducts. This conclusion was further supported by elevated rate of Pt-DNA adduct loss from the cisplatin-resistant tumor cells compared to the non-resistant lines [58–60]. DNA provides plentiful binding sites for Pt owing to its size and chemical complexity [61]. Moreover, although the kinetics and mechanisms of Pt(II) nucleophilic substitution of amine halides remains well understood, there was no prior reason to assume the involvement of this native behavior in the binding of cis- and transplatin to DNA.

7.2.2 Kinetics of DNA Binding Activities Pt NMR spectroscopy is normally used to study cis- and transplatin binding, to short duplex DNA fragments (30–50 base pairs, average molecular weight (MW) = 25,000) [62]. The reaction commences with Cl− replacement by water in a rate determining solvolysis step. This is followed by DNA binding to form monofunctional adducts, subsequently ensued on a rapid scale. Closure to form bi-functional adducts is kinetically regulated by second Cl− loss, for which estimated t1/2 values were 2.1 and 3.1 h, for cis- and transplatin, respectively, under identical conditions. The large chemical shift range for 195Pt imparts feasibility to follow these individual steps. 195

7.2.3 Structural and Regioselectivity of DNA Adducts Although the rate and mechanism of DNA adduct formation with cis- and transplatin do not differ significantly, the region and stereo specificity for the two complexes are convincingly distinct. In particular, cisplatin prefers to form 1,2-intrastrand cross-links between adjacent nucleotide bases, with 90% of the adducts having Pt coordinated to the N7 positions of two guanine bases or an adenine and guanine base. The structure of most

Platinum Complexes in Medicine  211 prevalent adducts were determined using X-ray crystallography [63]. The model building [64] and NMR [65, 66] studies of short oligonucleotide fragments, however, deciphered an inability of trans isomer to form analogous 1,2-intrastrand cross-link. Such a tendency fascinates a coordination chemist since this adduct involves a 17-membered chelate ring, alongside moderate stereochemical requirements of the DNA chain, making it feasible to accommodate the transpositions of transplatin moiety. Instead, the trans isomer forms a variety of DNA adducts [67, 68], including 1,3-­intrastrand cross-links, several of which are now synthesized and characterized using NMR spectroscopy. Two purine N7 atoms are linked by Pt to enclose a 23-membered ring. In one instance, involving the 12 nucleotide sequenced (TCTACGCG’ITCT), [the 1,3-intrastrand transplatin (N7-G(6),N7-G(8)) adduct] was noted as metastable, realigning with 47 h duration half-life at 37oC to provide an equilibrium mixture containing more stable, intrastrand N3-C(5), N7-G(8) crosslink [69]. For cisplatin, both linear and bent DNA molecules containing the cisplatin-d(GpG) adduct are known [70], but recent studies of duplex oligonucleotides containing this and the related cisplatin-d(ApG) adduct reveal the bent structure as the major product [71, 72]. For transplatin, stable duplex structure is reported, via molecular dynamics calculations. The two amine ligands form a hydrogen bonded bridge between the two DNA strands and the intervening nucleotide(s) stack into the minor groove. It has not been possible to estimate a bending angle for the transplatin adduct. Moreover, Pt appears as the central point of interest for a hinge joint rather than the induced cisplatin bend [73]. An important direction for the near future is to gather the experimental evidence for the actual adduct structures formed by cis and trans isomers with DNA, preferably using X-ray diffraction.

7.2.4 Studies on Action Mechanism The foremost domain in elucidating the cisplatin molecular mechanism is the evaluation of processing conditions corresponding to formation of specific DNA adducts under in vivo conditions and the correlation of gathered information with anticancer effects. The relative biological activities of different cisplatin-DNA adducts have, thus far been difficult to determine since most studies have been conducted using DNA containing all possible cisplatin adducts. It seems likely, however, that only the two major cisplatin adducts are instrumental in inhibiting DNA replication. The low level of bound drug required to inhibit DNA replication [for example, a platinated SV40 genome (50% inhibition at about 4 Pt per genome)] implies that low frequency adducts, such as inter and intrastrand cisplatin-d(GpNpG)

212  Advances in Metallodrugs crosslinks, should not be present on most SV40 molecules during the replication inhibition [74, 75]. Similarly, low levels of bound cisplatin also prevented the replication of plasmid-sized DNA in other systems [76], again inferring two major cisplatin adducts, [with d(GpG)] and d(ApG)] as substantial contributors towards DNA replication inhibition. Perhaps, there seems no validated reason a priori to expect identical biological activities from different cisplatin-DNA adducts. For example, different adducts are not equally mutagenic. Predominantly, single base substitution mutations are observed in E. coli, although frame shift mutations also prevail [77–80]. Several studies have demonstrated variable activities of different adducts, predicting cisplatin-d(GpNpG) or cisplatin-d(ApG) as the most mutagenic species. Recently, the combination of coordination chemistry and molecular biology has provided DNA molecules containing specific cis- and transplatin adducts [81–83], capable of being evaluated for their replicating/ repairable attributes, in vitro or in vivo. Engineering of specific cisplatin adducts into DNA in vitro should facilitate a more accurate comparison of cisplatin-DNA adduct biological activities. Moreover, such systems allow assessment of sans complication Pt binding to DNA, arising from Pt induced modification of other biomolecules [84–87]. In one experiment, a cisplatin-d(GpG) adduct was introduced into a specific site in the E. coli bacteriophage Ml3 1961 plus strand. The single Pt adduct reduced the transformation efficiency of the single-stranded phage DNA by 90%, compared to an unmodified Ml3 control, presumably via inhibition of first round DNA replication events. Similarly, cisplatin modification of purified SV40 DNA inhibits subsequent DNA replication in vitro, at biologically relevant drug-to-nucleotide levels [88]. Although the intrastrand crosslinks formed by cis- and transplatin substantially differ, both inhibit DNA replication in vitro and in vivo. The cis isomer is far more toxic and mutagenic to cells in culture than transplatin. In order to obtain equitoxic amounts of trans isomer bound DNA, far greater drug (here cisplatin) quantities must be added to the culture media. Two possible explanations have been suggested to account for this, with the first concluding that adducts formed in vivo by transplatin inhibit DNA replication less efficiently than those of cisplatin. Perhaps these adducts are monofunctional, since bifunctional adducts of the cis and trans isomers are equally effective in binding replication sites. Monofunctional transplatin-DNA adducts may react faster with cellular components, especially those containing sulfhydryl groups such as glutathione. Furthermore, from 195Pt NMR experiments, it is now well-known that glutathione reacts much rapidly with transplatin monofunctional adducts than those formed by cisplatin. Such reactions might deactivate transplatin through forbidding the

Platinum Complexes in Medicine  213 formation of toxic bifunction lesions on DNA. Alternatively, there may be more efficient repair of Pt-DNA adducts formed by the trans isomer, including possible cross-links between DNA and glutathione. Further studies of these possibilities are warranted. The repair of specific cisplatin adducts from DNA is also pertinent to be evaluated, since it plays a key role in conferring Pt cytotoxicity. In the best characterized example to date, purified components of the E. coli ABC exo-nuclease repair system efficiently eliminate the cisplatin-DNA adducts, in vitro [89, 90]. In this system, adducts are excised as 12 carbon comprising nucleotides following endonuclease cleavage of the damaged DNA strand eight nucleotides upstream (5’) and four nucleotides downstream (3’) from the site of Pt binding. The resulting gap is filled by DNA polymerase subsequent to sealing by DNA ligase. Once again, in a related series of experiments, specific DNA adducts were not found as being processed identically. In particular, [Pt(DACH){d(ApG)}] was repaired much more swiftly than [Pt(DACH) {d(GpG)}], and both were repaired less efficiently than [Pt(DACH){d(GpNpG)}] or monofunctional adducts (where DACH represents 1,2-­ diaminocyclohexane). The specific DNA distortions responsible for repair mechanisms are still poorly comprehended. Several efforts are being made to decisively isolate the factor(s) responsible for the recognition and subsequent repair of specific cisplatin-DNA adducts in eukaryotic cells. This objective has not been accomplished to date, although proteins that bind specifically to DNA were noticed as modified using cisplatin in cell extracts and cloned populations [91, 92]. Another question of scientific and clinical importance is the mechanism of acquired resistance towards cisplatin [93]. It is likely that several different mechanisms contribute, in light of unanimous reports pertaining to increased levels of metallothionein [94, 95], glutathione [96, 97], thymidylate synthase activity [98], and DNA repair in cisplatin-resistant cells. In the first two instances, the cells apparently utilize Pt affinity for sulfur donor ligands to bind and remove the toxic agent from the cytoplasm. Elevated drug efflux, a major resistance mechanism for most chemotherapeutic compounds [97], has not yet been identified in cisplatin-resistant cells. As far as DNA repair mechanism is concerned, the understanding of cisplatin resistance mechanisms may help coordination chemists in designing potential complexes, capable of resolving this issue, which otherwise, arrested clinical efficacy of drug(s). Although cisplatin inhibits DNA replication via blocking cell division, the exact mechanisms by which it kills the cells remain un-­understood. Apart from inhibiting replication, cisplatin also inhibits DNA transcription and amino acid transports, even though exact correlation of these suppressed activities and cytotoxicity remain unclear. As might be expected,

214  Advances in Metallodrugs the biochemical basis for selective cisplatin cytotoxicity towards certain cancer cell types is also unknown with underlying cause of Pt-induced cell death, still under investigation [98]. It seems reasonable to expect, however, that research into the mechanisms of cisplatin repair, resistance, and cytotoxicity will, eventually, aid in the ability of cisplatin chemotherapy to cure testicular cancer at minimal dose levels, causing little harm to normal cells. In nut shell, there is presently, much detailed information available about the coordination chemistry of Pt anticancer drugs and their dative analogs with DNA. The question still remaining unanswered is the distinctive evaluation of persistence and repair of individual adducts, in normal and tumor cells from a variety of tissues. Perhaps, the anticancer properties of cisplatin arise from the inability of tumor cells to repair lesions at a rate compatible to allow survival, whereas normal cells of the same type are more efficient at excising platinum injure. If such differential repair mechanisms were responsible for the chemotherapeutic properties of cisplatin, coordination chemists would have capitalized on a rational basis pertaining to the design of new and more potent metal-based anticancer drugs.

7.3 Pt Complexes as Anticancer Drugs Table 7.1 reports the most well-known conventional Pt anticancer complexes, with their utility in the treatment of various cancers. Initially, the cisplatin was wildly used to prevent cancer but the clinical limitations have led to the creation of thousands of cisplatin analogs, resulting in complexes that employ the same mechanism of action, through which Pt binds to DNA.

7.3.1 DNA-Coordinating Pt(II) Complexes While the conventional Pt(II) drugs mentioned so far exert their anticancer effect via coordinating, typically to the guanine DNA bases, there are many other DNA binding modes. These differing binding modes are or have been proposed, many a times to great effect, by various drugs and drug candidates. For example, groove binding facilitates reversible intermolecular associations where molecules could be designed to match the topologies of major or minor DNA grooves [100]. Such groove binders may be selectively designed to target specific DNA sequences, typically encompassing several base pairs. This type of binding induces only small changes to the overall DNA structure, inferred via viscosity and linear dichroism experiments. The binding of organic groove binders, such as distamycin and

Name of drug

Cisplatin

Oxaliplatin

Satraplatin

Sr. no.

1

2

3

Cl

Cl Pt

H2 N O Pt N2 O H

NH3

NH3

Structure

Pt

O

NH2 O

H3N

O

O

O

Cl

Cl

O

CH3

CH3

Table 7.1  Initially used Pt drugs: Cisplatin and its analogous.

(Continued)

Lung cancer, mesothelioma, brain tumors, and neuroblastoma

Colorectal cancer

Testicular, ovarian, cervical, breast, bladder, head and neck, oesophagal, lung cancer, mesothelioma, brain tumors, and neuroblastoma

Uses

Platinum Complexes in Medicine  215

Name of drug

Carboplatin

Nedaplatin

Heptaplatin

Sr. no.

4

5

6

O

H3N

H3N Pt

Structure

O

O

O

O

Pt NH3

NH3

O

O

NH2 O

Pt

NH2 O

O

O

Table 7.1  Initially used Pt drugs: Cisplatin and its analogous. (Continued)

Advanced gastric cancer

Non-small cell lung, small cell lung, oesophagal together with head and neck cancers

Ovarian cancer, lung cancer, head and neck cancer, brain cancer, and neuroblastoma

Uses

216  Advances in Metallodrugs

Platinum Complexes in Medicine  217 Hoechst 33258, has been suggested as being driven through a combination of hydrophobic, electrostatic, H-bonding, and van der Waals interactions. The preference of AT sequences in the minor groove of the DNA concerned is critical. Polyamides, N-methylpyrrole (Py), N-methylimidazole (Im), and N-methyl-3-hydroxypyrrole (Hp) form four ring-pairings (Im/Py, Py/Im, Hp/Py, Py/Hp), capable of recognizing the four base pairs of a fixed length [101, 102]. This sequence selectivity of groove binders has emerged as a boost for designing unique Pt(II) complexes [103]. Examples include: [Pt(distamycin)Cl2], [Pt(distamycin)2Cl2] [104], and trans-chlorodiamine[N-(6-aminoethyl)-4-[4-(Nmethylimidazole-2-­ carboxamido)-N-methylpyrrole-2-carboxamido]-N-methylpyrrole2-carboxamide] Pt(II)chloride (Figure 7.3) [105]. Eminent examples of polynuclear Pt(II) groove binders include [{trans-PtCl(NH 3) 2} (N 2H(CH 2) 4NH 2)] 2+ and BBR3564([trans-diamminechlorido Pt(II)] [(l-trans-diammine dihexane diamine-N,N) Pt(II)]. This approach stimulates the affinity for DNA [69, 70]. BBR3564 exhibits considerable in vitro and in vivo potency [106], forming DNA adducts that are unable of being recognized by the repair proteins [107–110]. The maximum tolerable dosage extents of these entities have been further restricted by sufferers in phase II clinical trials (exhibition of substantial neutropenia, gastrointestinal toxicity and other side effects) [111]. This has necessitated further studies to optimize the extent of such threats [112]. Intercalation is another mode of reversible intermolecular association, stabilized via simultaneous contributions of electrostatic, H-bonding, entropic, van der Waals, and hydrophobic interactions in the optimization

2+ H3N Pt Cl

H2 N NH3

H2 N H3N

Cl Pt

NH3 4+

H3N Cl

Pt

H2 N NH3

NH2 H3N

Pt

Cl NH2

NH2 H3N

NH3 Pt Cl

Figure 7.3  Examples of Pt complexes interacting DNA by groove and coordinate binding.

218  Advances in Metallodrugs of DNA targeting [113]. Here, a molecule, with an electron deficient planar aromatic ring system, inserts either entirely or partially between two adjacent DNA base pairs and is stabilized via π-π stacking. Intercalation results in unwinding, lengthening, and stiffening of the strand(s), resulting in the loss of regular DNA (helical) structure. The intensities of changes are subject to the depth of insertion [114–120]. Covalently binding Pt(II) complexes, incorporating an organic intercalator, such as phenanthroline, anthraquinone, phenanthridine, acridine, anthracene, or ellipticine along with a labile leaving group, often exhibits intercalation followed by coordinative binding with DNA [121]. Examples where the intercalating species is/are directly coordinated to Pt include: (terpyridine)Pt(II)chloride [Pt(terpy)Cl]+ and 2-hydroxyethanethiolato(terpyridine) Pt(II) and ([Pt(terpy)HET]2+, one of which is presented in Figure 7.4 [122, 123]. Complexes incorporate a tethered intercalator designed for providing stronger flexibility alongside facilitating coordination in forming monofunctional DNA adducts and evading normal DNA repair mechanisms [124]. The cytotoxicity of Pt(II) in H460 non-small-cell lung carcinoma stands comparable with cisplatin [125]. Further design modifications (as inferred from computational studies) are reported, with acridine replacement, either by benz[c]acridine (PtII-25) or 7-amino-benz[c]acridine (PtII), so as to obtain comparatively less cytotoxic complexes, although IC50 values were noticed in the micro molar range. A highly active series of tetraplatinated porphyrins have been reported using photodynamic therapy (PDT) agents, with cytotoxicity instigated through light irradiation [126]. The lead compound in this series (PtII-27, Figure 7.5) exhibited an IC50 value of 100 μM in CP70 cisplatin-resistant human ovarian cancer cells. 2+

N+

H2N

H2 N H3N

NH3 Pt Cl

Figure 7.4  Examples of Pt complex: direct intercalating coordination to Pt.

Platinum Complexes in Medicine  219 4+ NH2

H3N

Cl N

N

Cl

NH3 Pt

Pt

NH2

N H N

N H N

H2N

Pt

Cl

N

N

Pt Cl

H3N

H2N

NH3

Figure 7.5  Tetraplatinated porphyrins complex.

7.3.2 DNA-Covalently Binding Pt(II) Complexes Several non-covalent organic intercalators, such as doxorubicin [127], daunomycin [128], mitoxantrone [129], and amsacrine [130], are permitted by FDA for human cancer treatment (Figure 7.6). These moieties are known to disrupt DNA synthesis and repair via inducing reversible interactions with DNA base pairs along with having significant topoisomerase inhibition activity via inducing single- or double-stranded (topoisomerase I or II) breaks [131–133]. The inherent activities of these agents could be modulated by varying the size of planar aromatic area and the type or length of the side chains. A fine example is Mitoxantrone, which not only induces intercalation but also electrostatic interactions driven binding [134]. This design has been carried forward by mononuclear Pt(II) complexes to elicit the same mode of action via non-covalent DNA binding; resulting complexes through this modification demonstrated considerable activity, augmenting further interest in their investigation as conventional Pt complexes alternatives. Examples of intercalating square-planar Pt complexes are the compounds which incorporate heterocyclic polyaromatic ligands such as terpyridine and 1,10-phenanthroline [135]. X-ray structures have demonstrated that polyaromatic

220  Advances in Metallodrugs NH2

O

O

OH

O

OH

HO

OH

O

O

O

NH

HN OH O

OH

OH

O

OH

O

OH

O

OH

(c)

OH

(b)

NH2

(a)

O

HN

NH HO

O

O

OH

O

H N

O

S O

HN

N O

(d)

Figure 7.6  Non-covalent organic intercalators: (a) Doxorubicin, (b) Mitoxantrone, (c) Daunomycin, and (d) Amsacrine.

moiety inserts into DNA base pairs [136] along with cytotoxicity studies exhibited a wide range of activities [137]. A cyclo-metallated Pt(II) complex, [Pt(6phbpy)(tert-butylisocyanide)]+, (where 6phbpy is 6-phenyl2,20-­bipyridine) was reported to exhibit 0.009 and 0.01 μM, IC50 values towards human oral epidermal carcinoma and SH-5YSY neuroblastoma cells, respectively. Pt(II) stabilizes the topoisomerase-I–DNA complex, proposed to determine the inherent action mechanism. Subsequent in vivo testing on nude mice, implanted with NCI-H460 lung cancer, revealed 60% tumor inhibition with no significant side effects [138]. This collection of complexes displayed luminescent attributes, which increased upon intercalation [139]. Complexes incorporating 2-phenyl-6-(1H-pyrazol-3-yl)pyridine were reported to cause cell death via lysosomal accumulation. Recently, yet another group of organoplatinum complexes were also reported, with [Pt(R-dmba)LCl] like probable arrangement, where R could be −OMe, Me, H, Br, F, CF3, or NO2; dmba stands N,Ndimethylbenzylamine and L stands DMSO or tris(4-(trifluoromethyl)-­phenyl) phosphine (P(C6H4CF3-p)3) [140]. In vitro studies on ovarian (A2780) and cisplatin-resistant ovarian cancer (A2780cisR) cell lines exhibited 0.63 and 1.97 μM IC50, respectively, for the lead Pt(II) compound compared to cisplatin (1.90 and 19.57 μM). Accumulation studies revealed higher Pt(II)

Platinum Complexes in Medicine  221 concentrations than cisplatin in both cell lines, while cell cycle studies deciphered Pt(II) mediated cell-cycle arrest in the G0/G1 phase, contrary to cisplatin where the cell cycle arrest happens in S-phase. Both these factors seem critical for the observed differences in cell-killing potency [141]. [Pt(1-(8-quinolylamino)-3-(8-quinolylimino)-propene)]+ Pt(II) is another complex exhibiting emission when intercalated into DNA base pairs as well as considerable in vitro anticancer activity (equal or greater than cisplatin) in A2780 and A2780CP70 cell lines, having 3.2 ± 0.5 and 3.7 ± 0.1 μM IC50 values. A comprehensive series of [Pt(PL)(AL)]2+-type complexes are also reported (where PL signifies a polyaromatic ligand) [142] [including 1,10-phenanthroline (phen), 5-methyl-1,10-phenanthroline (5-Mephen), 5,6-dimethyl-1,10-phenanthroline (56Me2phen, Pt(II)-31)), 2,20-bipyridine (bpy),4,40-dimethyl-bpy (44Me2bpy), dipyrido[3,2-f:20,30-h] quinoxaline (dpq), or 2,3-dimethyl-dpq (23Me2dpq)] and AL is an ancillary bidentate ligand which can be achiral [1,2-diamino ethane (en) [65], 1,3-diaminopropane (1,3-pn), 1,3-diamino-2-hydroxy propane (1,3-pnOH), 1,4-diamino butane (1,4-bn) or cis-1,4-diamino cyclohexane (1,4-dach)] or chiral [1,2 R-diamino propane (1,2R-pn)], 1, 2S-diamino propane (1,2S-pn), 1R,3S-diamino-1,2,2-trimethyl cyclopentane (1R,3S-tmcp), 1S,3R diamino-1,2,2-trimethyl cyclopentane (1S,3Rtmcp), 2R,3R diaminobutane (R,R-bn), 2S,3S-diamino butane (S,S-bn), 1R,2R diamino cyclopentane (R,R-dacp), 1S,2S-diamino cyclopentane (S,Sdacp), 1R,2R-diaminocyclohexane (R,R-dach), 1S,2S diaminocyclohexane (S,S-dach), N,N0-dimethyl-1R,2Rdiaminocyclohexane (NMe2-R, R-dach), N,N0-dimethyl-1S,2Sdiaminocyclohexane (NMe2-S,S-dach), N-benzyl-1S,2Sdiaminocyclohexane (NBn-S,S-dach), 1R,3S-diamino-1,2,2trimethylcyclopentane (R,S-tmcp) and 1S,3R-diamino-1,2,2-trimethyl cyclopentane (S,R-tmcp), 1S,2S-diphenyldiaminoethane (S,S-dpen), 1R,2Rdiphenyldiaminoethane (R,R-dpen), rac-1,2-bis(4-­ f luorophenyl)-1,2diaminoethane (F2phen) or rac-1,2-diamino-­4methyl-cyclohexane (4Medach) [143–154]. One of these complexes is depicted in Figure 7.7. Major factors contributing to cytotoxicity include charge regulated solubility (positive charge for a higher value), effective cellular uptake via active transport, and high DNA affinity. Although the mechanism has not been elucidated, yet it is profoundly distinct from cisplatin [134, 155]. Despite its feasibility, intercalation alone does not illustrate the activity differences of (cf 56MESS (Pt(II)-31) and (56MERR). A more considerable influence on cytotoxicity results from AL changes, complimenting the conclusion that DNA binding as not the only factor responsible for it. Studies have suggested that 56MESS acts at mitochondrial level, enhancing the protein MTC02 expression [156]. Complementary

222  Advances in Metallodrugs 2+ H2 N

N Pt N

N H2

Figure 7.7  Pt complex of the type [Pt(PL)(AL)]2+.

DNA microarrays, used to measure the genome-wide gene expression changes in Saccharomyces cerevisiae revealed an involvement of 56MESS up-regulated genes in restoring cellular homeostasis towards Cu and Fe transport [157]. It was subsequently reasoned that the reduction of intracellular Cu and Fe transport by 56MESS activated the regulation genes through the involvement of transcription factors. This interpretation was further augmented by diminished Cu and Fe levels in 56MESS treated cells alongside high-affinity 56MESS sensitivity of Fe transporter Ftr1p mutants [158–160]. 56MESS (Pt(II)) down-regulates the methionine, cysteine, and the peptide transporter gene PTR2 biosynthesis, deciphering insignificant expression of glutathione-mediated resistance [161]. Although it induced the simultaneous down-regulation of several genes regulating cellular defenses, none relates to DNA repair mechanisms. This observation further suggests that the cytotoxicity of 56MESS is the manifestation of manifold disrupted cellular defense mechanisms, and not by the as expected, DNA binding. 56MESS [119] exhibits the nanomolar IC50 in many cancer cells [128], including L1210 cell line, where it is 100-fold superior to cisplatin [120]. 56MESS remains well-known for its superior activity in cisplatin-resistant cell lines [114], but the in vivo studies could retrieve only low activity accompanied with dose-limiting nephrotoxicity [139]. One strategy currently being pursued to overcome these hurdles is to design Pt(IV) derivatives, using axial ligands, supposedly with improved pharmacological properties [62, 162].

7.3.3 Targeted Pt(II) Complexes In the quest for drugs that functions similar to Ehrlich’s “magic bullet”, researchers have employed a strategy that combines proven anticancer Pt(II) complexes with targeting molecules to selectively deliver cytotoxic

Platinum Complexes in Medicine  223 payloads to cancer cells [163]. An appropriate targeting vector is one that is overexpressed on the cancer cell surface [164] or the one, which can be directed to specific organelles at the sub-cellular level to develop cytotoxicity. This strategy can also be used to direct Pt(II) complexes to cancerous tissues, meanwhile looking for proteins that are expressed on angiogenic blood vessels or by utilizing molecules that are activated via acidic or hypoxic microenvironment within a cancerous mass [165]. Overexpression of glucose membrane transporters remains a characteristic feature of cancer cells as increased energy (input) is required for cell division, a prospect exploited with the use of 18F 2-fluoro-2-deoxy-D-­ glucose in positron emission tomography (PET) imaging [60, 166]. Several derivatives of Pt(II)-sugar complexes have been studied to capitalize this attribute through a higher water solubility as well as enhanced cellular uptake (for glucose transporters). Complexes of many kinds and variable chirality and proximity (of the coordinated sugar moiety) have been evaluated for this purpose [167]. For D- and L-glucose conjugates, naturally occurring D-isomer is reported more commonly, suggesting a likelihood involvement of a specific receptor, although no evidence regarding this was established [168]. Confirmation regarding Pt(II)-sugar complexes interaction with the glucose receptor was acquired through in vitro cytotoxicity assays, where block receptors exhibited decreased [Pt(R,R-dach) (2-halidomalonate sugar)] binding efficacy towards Pt(II) (Figure 7.8) [169]. Modifications to this design were retrieved following the attachment at 6th position, conferring benefit from the D-glucose selectivity, since the –OH group does not interact with protein side chains [170]. The resulting complex, [Pt(R,R-dach)(malonate-X-6-sugar)] (Pt(II), (Figure 7.8) with variable linker lengths (X), exhibited a selective cancer cell uptake. Further experiments confirmed that different glucose transport inhibitors together with organic cation transporters contributed to uptake efficacy [171]. Interest in oestrogen receptor (ER), as the target molecule for Pt(II) complexes gained prominence after its documentation as being overexpressed on the surface of multiple cancers, with a sharp influence in breast cancer, where it encourages cell proliferation [60]. A number of ER targeted Pt(II) OH

OH O

HO

O

X

Pt

O

HO OH

NH2

O

Figure 7.8  Pt(II)-sugar.

N H2

O

HO

O

HO

H3C O

OH

NH2 Pt

O

N H2

224  Advances in Metallodrugs complexes have been prepared where the oestrogen resides at some distance from the Pt, so as to not impede the interaction with ER, Pt(II), and Pt(II) [172]. Another molecules finely fitted into this domain are glycoproteins, overexpressed on the surface of many tumor cells and function as integrated folic acid receptors (FR), presenting interest as effective Pt complexes targets [173]. There have been fewer examples using this approach, with cisplatin and carboplatin complexes [Pt(II), Figure 7.8] carrying folate moiety, exhibiting a poor water solubility [174]. While the water solubility is enhanced via introduction of a PEG spacer (Pt(II), Figure 7.10), the complex was noted as less cytotoxic, forming fewer DNA adducts compared to carboplatin. The CD13 receptor, overexpressed on prostate cancer cells, was successfully targeted by a Pt(II)-PEGylated cyclic peptide complex Pt(II). The improved cytotoxicity, compared to carboplatin, attributed to a higher CD13 affinity [175]. Sub-cellular targeting was achieved on incorporation of a mitochondrial penetrating decapeptide [Pt(II), Figure 7.10]. The prevalence of Pt(II)-decapeptide conjugate in the ovarian cancer cell mitochondria was confirmed through fluorescence microscopy alongside preferential platination of mitochondrial DNA [176].

7.3.4 Pt(IV) Prodrugs The development of Pt(IV) prodrugs is an increasingly attractive field as this oxidation state provides many advantages over Pt(II), in in vivo conditions. Upon oxidation to Pt(IV), the complexes convert from four-­ coordinate square planar to six-coordinate octahedral, geometry, with the addition of two ligands in the axial positions. In general, Pt(IV) complexes are kinetically more inert than the parent Pt(II) complexes, enabling their much simpler oral administration [62, 177]. Once inside cancer cells, an ideal Pt(IV) complex will undergo activation by a two-electron reduction with simultaneous release of parent cytotoxic Pt(II) drug as well as the two axial ligands [178]. Several Pt(IV) analogs of cisplatin, iproplatin (ctcdichloridodihydroxidobis (isopropylamine) Pt(IV), tetraplatin (ctc-­ tetrachlorido(1,2-cyclohexanediamine-N,N0)Pt(IV)], ormaplatin, PtIV-2), and satraplatin (bis(acetato-O) ammine dichlorido(cyclohexylamine) Pt(IV), Pt(IV-3), (Figure 7.9), have been assessed in clinical trials, unfortunately none have been approved for clinical use. Iproplatin demonstrated a lower activity compared to cisplatin while tetraplatin exhibited a higher neurotoxicity [179, 180]. Likewise, Satraplatin can be orally administered but is not FDA approved since it did not significantly improve the overall survival compared to cisplatin reference standard [62, 181, 182]. Results of these first generation Pt(IV) complexes, with

Platinum Complexes in Medicine  225 O

H3N

Cl H2 N N H2

Cl Cl

HO

Cl

H3N

Cl

CH3

Pt

NH2 O

Pt

Pt Cl

OH

O

Cl

CH3

NH3 Cl

O

Figure 7.9  Structure of Tetraplatin, Oxoplatin, Satraplatin, respectively: Pt (IV) complex.

simple conjugated axial ligands such as hydroxido, chloro, or acetate, have inspired further innovations in the choice of axial ligands, emerging critical for regulating pharmacological properties. These comprised enhanced lipophilicity, increased solubility, modified reduction rates, targeting vectors, and incorporation of additional bioactive molecules [183, 184].

7.3.5 Multiple Action of Pt(IV) Prodrugs Manifold action potential of Pt(IV) derivatives can be considered through presuming them as combination therapy variants, characterized by the two anti-proliferative axial ligands, which once reduced, are released inside the cancer cell and allow action at different cellular targets via significantly unrelated mechanisms. This increased the probability of overcoming the drug resistance, in a single mode of drug activity. Multiple action prodrugs have the distinct advantage of presenting a single pharmacokinetic profile, effectively overcoming cisplatin resistance [148, 152]. Significant to cisplatin resistance is the nucleotide excision repair (NER) pathway, responsible for the repair of covalent DNA lesions, such as intra-strand cross-links. Recently, a significant NER inhibitor, (E)-2-(((8a,9a-dihydro9Hfluoren-9-ylidene) hydrazono)methyl) benzoic acid, was conjugated at the axial position of a Pt(IV) cisplatin analog, to produce NERi-Pt(IV) (Figure 7.12). This complex exhibited increased activity against cisplatin resistant ovarian (A2780cisR) and non-small-cell lung cancers (A549cisR) with 0.300 and 0.290 mM, IC50 values. Mechanistic studies on A549 and A549cisR cells revealed higher intracellular accumulation in cells together with Pt-GG intra-strand cross-links compared to cisplatin. Increased cytotoxicity was attributed to NER inhibition. Combination therapy, incorporating Pt(II) anticancer drugs with histone deacetylases inhibitors

226  Advances in Metallodrugs (HDACi) such as dichloroacetate (DCA), valproate (VPA), and phenyl butyrate (PhB), together comprise an emerging class of anticancer drugs. HDAC confers an altered gene transcription affecting cell-cycle arrest, differentiation, apoptosis, and tumor angiogenesis inhibition, with improved patient outcomes [175–185]. Dichloroacetate inhibits pyruvate dehydrogenase kinase (PDK), shifting cellular metabolism away from glycolysis (reversing the Warburg effect), alongside restoring normal mitochondrial function [125]. Mitaplatin is a Pt(IV) derivative of cisplatin with two bioactive axial dichloroacetato (DCA) ligands [Pt(IV), Figure 7.12]. Pt(IV) is cytotoxic and has a cisplatin comparable cytotoxicity in various cancer cell lines. Upon reduction, the dual action proceeds through DNA involving covalent interaction and an attenuated mitochondrial activity via altering the DCA regulated mitochondrial membrane potential gradient of cancer cells. Reported to kill cancer cells selectively, Mitaplatin is capable of partially overcoming the cisplatin resistance, although the Pt(IV) oxaliplatin derivative with DCA [Pt(IV), Figure 7.9) does not exhibit an improved cytotoxicity. The Pt(IV) dual action prodrugs, capable of combining cisplatin with VPA [Pt(IV)] or PhB [Pt(IV)] (Figure 7.10), have been studied to exhibit nearly 100-fold higher potencies than cisplatin in many human cell lines [175], with the exception of Pt(IV) and much less potent higher grade oxaliplatin derivatives. Ethacraplatin [Pt(IV)] contains cisplatin and two axially coordinated etacrynic acid [2-(2,3-dichloro-4-(2-­ methylenebutanoyl) phenoxy) acetic acid] ligands, capable of inhibiting glutathione S-transferase (GST), consequently interfering in cisplatin detoxification [86]. Once in the cell, cisplatin implicitly interacts with DNA to form adducts while etacrynic acid (inhibiting GST), reversed the cisplatin resistance and impeded the resistant expression in breast, lung, and colon cancers. Chalcones, either as natural compounds or synthetic derivatives, have also been significantly reported for anticancer activities. They bind to the

O H2N H2N

O Pt OH

O Cl Cl

N

H2N H2N

O Pt O O

Cl Cl

H2N H2N

Cl Cl Cl

Figure 7.10  Structure of Pt (IV) complex.

O

O

Cl

O Pt O O

Cl

H2N

Cl

H2N

O Pt O O

Cl Cl

Platinum Complexes in Medicine  227 tumor suppressor p53 locus and simultaneously act as selective MDM2 antagonists, allowing p53 to remain active through obstructing the unchecked tumor proliferation [186]. Chalcoplatin [Pt(IV), Figure 10], a product of a Pt(IV) cisplatin core, having two axially coordinated chalcones, exhibits enhanced cytotoxicity, compared to cisplatin and on being administered individually or in combination with chalcones in several cell lines, such as cervical (HeLa), colorectal (HCT-116), breast (MCF-7), and lung (A549) cancer A549, which are all p53 wild-type cell lines [180]. Experimental evidence suggests this cytotoxicity as p53-dependent and being enhanced in the ­cisplatin-resistant A549 cell line. Endothall (3,6-endoxohexahydrophthalic acid) inhibits phosphatase 2A protein, responsible for phosphorylation and dephosphorylation. Coordination of two axial endothall ligands to a cisplatin scaffold Pt(IV) and Pt(IV) generates a dual action complex, not as cytotoxic as cisplatin [184]. The Pt(IV) complex ctc-[Pt(1S,2S-­diaminocyclohexane)(OH)2(5,6dimethyl 1,10-phenantroline)]2+ [56MESS(IV), Pt(IV), Figure  7.11] retained the cytotoxicity of “parent” Pt(II) complex 56MESS(II) across various cell lines, including that of prostate (Du145, 9 nM) and colon cancers (HT29, 22 nM) cisplatin at 1,200 and 11,300 nM, respectively [199]. Novel Pt(IV) 56MESS(IV) derivatives, comprising [acetate (OAc), octanoate (Oct), palmitate (Pal)] and/or bioactiveaxial ligands, such as PhB or VPA produce complexes that are significantly more potent than cisplatin [141, 186]. O

R2 R1

N

N

O Pt

Pt R2

R2

N

N

O

H3N

H2N

H2N

R1

H2N

H2N

H3N H N

O

R1

O Cl O

O Cl

O

N

O

H2N

Pt

H3N

H2N

N

H3N H N

O O

Figure 7.11  Chemical structure of 56MESS(IV) and its derivatives.

O Pt

Cl Cl

O O

O

Cl

Pt O

Cl

O

228  Advances in Metallodrugs Complexes of Pt(IV) exhibited 0.44 and 0.69 μM IC50 values against pancreatic adenocarcinoma (BxPC3) cell lines, respectively, considerably lower than 56MESS (Pt(II)), cisplatin and oxaliplatin. Cellular accumulation did not better with hydrophobically sensitive axial substituents, suggesting a role of mechanisms different from those of passive diffusion. Suppression of HDAC was the main mechanism for Pt(IV), presumably being expressed at 15 times higher than cisplatin. In vivo activity for C57BL mice bearing Lewis lung carcinoma treated with 56MESS(IV), Pt(IV), or cisplatin exhibited 61%, 73%, and 75% inhibitions, respectively. With comparable effectiveness, Pt(IV)-21 exhibited reasonable systemic toxicity, while treatment with cisplatin resulted in a more significant body weight reduction [141]. On this basis, dinuclear dual Pt(IV) and multiple action Pt(IV) prodrugs are being prepared. Pt(IV) combined cisplatin and 56MESS Pt(II), exhibited an enhanced activity in some cell lines (HCT-15, A375, and PSN1), providing a reasonable comparison with Pt(IV). Multiple actions were achieved through coordinating four different bioactive moieties, namely, cisplatin, DCA, PhB and Pt(II) (Figure 7.11). Pt(IV) produces covalent modification of DNA due to cisplatin, inhibits HDAC activity by PhB, and interferes with mitochondrial activity, supposedly due to the combined DCA and 56MESS Pt(II) activities [143]. Acting as a multiple active prodrug, Pt(IV) demonstrated significant potency compared to Pt(IV-25), cisplatin, oxaliplatin, and 56MESS Pt(II) in both 2D and 3D cell proliferation assays. Surprisingly, Pt(IV) demonstrated selectivity and potency against KRAS mutated cells, noteworthy because these cells are characterized by increased tumorigenicity and poor prognosis. Such a selectively distinct response could be due to RAS proteins participation in regulating cell differentiation, proliferation and survival, leading to being considered ineligible for drug formation [186].

7.3.6 Targeted Pt(IV) Prodrugs Effective targeting of cancer cells necessitates a distinctive recognition of differences in biochemical metabolism between “normal” and cancerous cells, exploited with a rationale of reducing toxicity and increasing efficacy. Critical to the effectiveness of this strategy are the selectivity, the Pt(IV) prodrug content being delivered to cancerous cells, and the potency of effective Pt complex [60]. Oestrogen receptors (ER), overexpressed on the surface of several cancer cells including uterine, breast, and ovarian, are known to induce upregulation of high-mobility group box 1 gene (HMGB1) which can avert DNA-cisplatin adduct repair [181]. A Pt(IV)

Platinum Complexes in Medicine  229 targeted prodrug, (Figure 7.12) facilitated the intracellular release of both cisplatin and oestrogen, bettering the cisplatin response [181]. A series of Pt(IV) cisplatin analogs incorporating peptide sequences (Figure 7.12) have been synthesized having selective ability of targeting formyl peptide receptors, FPR1 and 2, that are overexpressed in many metastatic cancers and immune cells. It was hypothesized that these Pt(IV) complexes operated through combined chemotherapy and immunotherapy mechanisms, specifically binding to immune cells for expression of a desired immune response. In vitro assessment of this efficacy against the three cell lines revealed FPR1 and 2 overexpression in malignant human glioblastoma (U-87MG), human breast cancer (MCF-7), and p53 mutant human breast cancer (MDA-MB-231). The most potent Pt(IV) complexes, reported till date, are the ones conjugated to protein labeled WKYMVm. Compared to cisplatin, a modest increase in anticancer activity was noted against U-87MG and MCF-7 cell lines (suggested through improved active targeting), although in MDA-MB-231 cell line, the activity was lower. Conversely, the fMLFK conjugate was effectively non-cytotoxic, suggesting the importance of peptide targeted sequence for receptor specificity. Pt(IV) derivatives of cisplatin and oxaliplatin, incorporating maleimide- and succinimide functionalized axial ligands, are reported to selectively bind serum albumin in the bloodstream [156]. This binding slowed the renal clearance of these complexes, which alongside paved way for a concomitant elevation in accumulating tumor tissues, facilitated through enhanced permeation and retention (EPR) effect. In vivo efficacy of these Pt(IV) complexes (PtIV-29-31) were assessed against BALB/c mice, bearing colon carcinoma (CT-26). Pt(IV)-29 displayed cisplatin comparable and even improved activity with respect to oxaliplatin. In an interesting attempt, Pt(IV)-31 incorporating an acetato axial ligand exhibited significantly O

O

O NH H3N H3N

O Pt

Cl OH

O H2N N H2

O

N

NH O

Cl

O

O

N

Pt

O O

O

O

R

Figure 7.12  Pt(IV) of Cisplatin analogs incorporating peptide sequences.

O

230  Advances in Metallodrugs higher activity than the methoxido conjugated Pt(IV)-30. While Pt(IV)31 cured 4 out of 8 mice bearing CT-26 colon carcinoma, the oxaliplatin exhibited moderate to severe toxicities [169]. Distribution studies on Pt complexes delivered for cancer treatment revealed higher cisplatin and oxaliplatin segregations in kidney and liver, compared to the tumor cells. Conversely, higher Pt(IV)-31 levels were noticed in tumor tissue, yet again because of the EPR effect. Targeting specific cellular differences, such as overexpression of folate receptor (FR) proteins on many cancer cell surfaces including those of lung, renal, endometrial, colon, ovarian, and breast, may be achieved through creating folate-drug conjugates, which in the meanwhile, facilitated an endocytosis mediated preferential cancer cell uptake [138, 159]. Another striking difference is that many cancer cells produce energy via glycolysis, to nearly 200 times higher extent than the “normal” cells (phenomenon, better recognized as Warburg effect). As glycolysis is expressed differentially, it is also recognized as a selective cellular target, in all tumor cells. One promising targeting vector being investigated in this context is prostate specific membrane antigen (PSMA), studied for fixing imaging agents and delivering the radiotherapeutics to 90% cells in prostate tumors. Functionalized soluble single-walled carbon nanotubes (SWNTs) have also been investigated for Pt(IV) delivery mediated cisplatin release [165–180]. This module comprises of a SWNT tethered Pt(IV) complex ([Pt(NH3)2Cl2(OEt)(O2CCH2CH2CO2H)]) [154], through one of its axial ligands to an amine functional group on its surface, which can deliver 65 Pt(IV) prodrugs per SWNT (bound), on an average. This design was further adapted to specifically target folic acid receptors on the cancer cells [186]. The resulting Pt(IV) complex, cct-[Pt(NH3)2])2Cl2 (O2CCH2CH2CO2H)(O2CCH2CH2CONH-PEG-folic acid), carried Pt(IV), coordinated succinate to one axial position facilitating the attachment of the amine-functionalized SWNT, while the other axial position is coordinated to a folic acid derivative. Solubility and biocompatibility of these Pt complexes are dependent on the separation of the centrally positioned Pt and folic acid receptor via incorporation of a PEG spacer [186]. One application for targeting prostate cancer utilized functionalized nanoparticles (NPs) developed from poly(D,L-lactic-coglycolic acid) (PLGA) and PEG to coordinate a hydrophobic Pt(IV) prodrug towards prostate specific membrane antigen (PSMA) [173, 179]. In this complex, the two linear hexyl chains regulated the drug release from the NPs to within the cancer cell [185]. Once inside the cell via endocytosis, the Pt(IV) is reduced to cisplatin. The Pt(IV) PLGA-PEG polymer is a safe administration tool, with moderate systemic clearance times [125].

Platinum Complexes in Medicine  231 H3C

N

Pt

N

Cl

Figure 7.13  Pt (II) complex a photodynamic killer of cancer cell.

7.3.7 Photodynamic Killing of Cancer Cell by Pt Complexes Recently, a new approach towards prevention of cancer has come out which is termed as “Photodynamic Therapy” [103]. It uses photosensitizers, those used in conjunction with molecular oxygen to elicit cell death. Upon light-­irradiation, these photosensitizers provide a strong alternative to conventional cancer treatment due to their ability to selectively target tumor material without affecting healthy tissue. Transition metal complexes are highly promising photodynamic therapy agents due to intense visible light absorption, yet the majority is toxic even without light. Such work has been done on platinum complex; Pt(2,6-dipyrido-4-Me-benzene) Cl (Figure 7.13) where this complex has found much effective in treatment of cancer.

7.4 Conclusion We have provided some insights on the anticancer attributes of Pt complexes, including a wide range of Pt(II) and Pt(IV) complexes. Several design strategies have been identified, including those emulating the coordinative/covalent structure and cisplatin action mechanisms (classical) as well as those which diverge both in structure and mechanism (unconventional or non-classical). The most interesting new Pt complexes are those which exhibit potent cytotoxicity in a manner significantly distinct from those used in the clinic. The transition to Pt(IV) scaffolds is swiftly emerging as the next logical step allowing the design flexibility that previously remained unachievable. Through a judicious selection of axial ligands liberated upon reduction, the physicochemical properties can be

232  Advances in Metallodrugs tuned for deploying additional bioactive axial ligands or selective incorporation of targeting moieties. The manifold impressive results demonstrated in the literature provide both, incentive as well as inspiration for the future potential of Pt-based anticancer therapies.

Acknowledgments The authors are grateful to Central University of Gujarat for providing infrastructure facilities round the clock internet facility.

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8 Recent Advances in Gold Complexes as Anticancer Agents Mohammad Nadeem Lone1*, Zubaid-ul-khazir2, Ghulam Nabi Yatoo2, Javid A. Banday2 and Irshad A. Wani3 Department of Biotechnology, School of Life Science, Central University of Kashmir, Ganderbal, India 2 Department of Chemistry, National Institute of Technology, Hazratbal, Srinagar, India 3 Department of Chemistry, Govt. Degree College, Anantnag, India

1

Abstract

The exploration of the mysterious anticancer properties of cisplatin by Barnett Rosenberg sparked a huge interest in exploiting the rich potential of metal complexes for the treatment of cancer. As a result, second and third generations of cisplatin analogs were developed with the claim of good anticancer properties and reduced side effects. However, persistence of some side effects and the resistance phenomena by cancer cells tempted scientists to explore new metal complexes as anticancer drugs and many such compounds have been developed and screened for various types of cancers. In view of these facts, evolution of metallo anticancer drugs has been discussed. More specifically, attempts have been made to describe the state-of-art of Au complexes as anticancer drugs. Besides, efforts have also been made to discuss the efficacy of Au complexes in nano-formulations. Lastly, future challenges and perspectives of Au anticancer drugs have been discussed. Keywords:  Gold complexes as anticancer agents, nano-formulations, future challenges, perspectives

*Corresponding author: [email protected]; [email protected] Shahid-ul-Islam, Athar Adil Hashmi and Salman Ahmad Khan (eds.) Advances in Metallodrugs: Preparation and Applications in Medicinal Chemistry, (247–272) © 2020 Scrivener Publishing LLC

247

248  Advances in Metallodrugs

8.1 Introduction Cancer stands as the second most common disease after cardiovascular responsible for many deaths all over the world [1]. Obviously, it has become a havoc for human beings with prediction about 11.0 million deaths globally in 2030 [2]. In 2016, there were 15.5 million (approx.) cancer survivors in the United States and the number of cancer survivors is expected to increase to 20.3 million by 2026. Besides, 15,270 children and adolescents (age: 0 to 19) were diagnosed with cancer. Out of this number, 1,790 cases died of the disease in 2017 [3]. Among the various types of cancers, the incidence of lung cancer in men was reported highest in the United States. Also the highest lung cancer incidence rates in women are reported in North America and parts of Europe including United Kingdom and Denmark [4]. The GLOBOCAN 2018 estimation reported 18.1 million new cases of cancer and 9.6 million deaths in 2018 [5]. Cancer is also the second most common disease in India, causing maximum number of deaths yearly [6]. The number of cancer patients is believed to increase in the developing and under developed countries in the coming decade to about 70%. Cancer havoc in the Indian sub-continent is increasing quite alarmingly due to poor and moderate living standards of the people [7] and insufficient medical facilities. Indian population mostly suffers from lung, breast, colon, rectum, stomach, and liver cancers [8]. Medicinal inorganic chemistry has brought about a grand revolution in the design of therapeutic agents with exciting properties, which are not possible with organic compounds [9]. The wide diversity of coordination numbers, geometries, redox states, thermodynamics, kinetics, and intrinsic properties of metal ion/atom and ligand offers a large variety of features to be exploited by a medicinal chemist [10]. Of course, cisplatin is the most famous metal complex used in the treatment of various cancerous malignancies and stands currently as one of the best-selling anticancer drugs worldwide [11]. Accidental discovery of the antitumor properties of cisplatin [12] was followed by one of the most exciting drug success stories ever; bringing about a significant improvement in cancer chemotherapy. Besides cisplatin, several other metal complexes have been approved for current cancer chemotherapy. However, only a limited number of tumors have been treated with these platinum-based anticancer drugs and the patients suffer from some deleterious side effects and it is the drug-­ resistance phenomenon that lowers the essentiality of these drugs [13, 14]. These drawbacks in platinum-based anticancer therapy tempted scientists towards the design of safe and effective non-platinum metal complexes as

Gold Complexes as Anticancer Agents  249 anticancer agents. Therefore, some metal ions such as copper, ruthenium, gold, palladium, iron, cobalt, titanium, gallium, and others have been exploited for this purpose. The interesting preclinical and clinical results of the above-mentioned metal complexes created a hope for human friendly medication [13]. This book chapter is confined to the gold complexes as anticancer agents. So, a thorough search of literature was carried out through Pubmed and ScienceDirect and it was found that more than 192 papers have been published on coordination complexes with gold centers as anticancer agents from 2014 onwards. The account of the number of papers published from 2014–2019 on gold and nano gold complexes as anticancer agents is depicted in Figure 8.1. The number of annually appearing papers on this topic has been increasing steadily. The present book chapter highlights the advances in the development of all gold complexes for the treatment of cancer till date. Besides, attempts have been made to discuss nano-­ formulations of gold complexes as chemotherapeutic modalities. Finally, the current challenges faced by inorganic medicinal chemists have been discussed with special emphasis on future perspectives of gold complexes as anticancer drugs. Hopefully, this book chapter will turn into a useful entity for the researchers actively involved in the design and development of gold anticancer drugs.

50 h searc in Re t s e r Inte asing Incre

45

Number of Research Papers

40 35 30 25 20 15 10 5 0 2014

2015

2016

Au Complexes

2017

2018

2019

Nano Au Complexes

Figure 8.1  A graphic representation of the increasing interest in research on gold complexes as anticancer agents from 2014 to 2019.

250  Advances in Metallodrugs

8.2 Evolution of Metal Complexes as Anticancer Agents Peyrone brought the revolution in cancer chemotherapy by synthesizing cisplatin in 1845. After a gap of more than hundred years, its biological activity was accidentally explored by Rosenberg and co-workers in 1965 [12, 15]. Rosenberg used inert platinum electrodes during the investigation of the influence of weak alternating currents on the growth of Escherichia coli. Researchers detected the inhibition of cell reproduction without simultaneous inhibition of bacterial growth, which eventually led to the formation of long, filamentous cells. This interesting detection was followed by extensive investigation, and it was concluded that lesser amount of platinum from the electrodes had reacted with ammonium chloride for the production of different platinum amine halide complexes. Among the formed complexes, cisplatin and its other tetrachloro complexes like cis-­tetrachlorodiamine platinum (IV) were capable to induce filamentous growth in the absence of electric field [12]. Later, Rosenberg et al. carried out the experiments on mice with sarcoma 180 and leukemia L1210 [16]. This led cisplatin to enter into phase I clinical trials in 1971; and in 1978, it was approved for the treatment of testicular and ovarian cancers. Nowadays, cisplatin is one of the most widely used antitumor drugs highly effective in treating testicular, ovarian, bladder, cervical, head, and neck and small-cell and non-small cell lung cancers (SCLC and NSCLC) [17, 18]. Despite high clinical success of cisplatin as an anticancer drug, it lacks tumor tissue selectivity leading to severe side effects like renal impairment, neurotoxicity, and ototoxicity (loss of balance/hearing), which are only partially reversible when the treatment is stopped. With a long-term or high-dose therapy of cisplatin, severe anaemia may develop in patients. To combat these problems, second and third generation ­platinum-based drugs were developed. Out of those, some were approved for the treatment of some cancers, few are in clinical trials and few complexes have not yet showed significant advantages over cisplatin [19]. The design of metal-based anticancer drugs was initially limited to platinum drugs only. Unfortunately, only few cancers could be treated with these drugs and the patients suffered from several side effects. Besides, the impacts of these drugs were further lowered by the drug-resistance phenomena [13]. These drawbacks of platinum drugbased chemotherapy stimulated research on other antitumor therapeutics with different metal ions such as copper, ruthenium, gold, palladium, iron, cobalt, gallium, nickel and others. Presently, considerable progress has been made in the development of other metal complexes as anticancer agents. Besides, the advent of nanotechnology has enabled the trapping of other metal anticancer complexes in nano identities for the safe and

Gold Complexes as Anticancer Agents  251

Non-Gold Complexes

Nano Structured Metal Complexes

Vanadium Zinc Cobalt

Gold Complexes

Miscellaneous Complexes

Nickel

Titanium Ruthenium Cis-platin

Copper

Silver

Iron Osmium

Iridium

Rhodium Molybdenum Metal Based Complexes

Figure 8.2  The development of metallodrug tree.

effective treatment of cancer. An overview of the evolution of anticancer metallodrugs is presented in Figure 8.2.

8.3 Gold Complexes Medical importance of gold has been identified thousands of years ago. However, it was not used in medicine until the early 1920s. Gold(III) is isoelectronic with platinum(II). Besides, tetracoordinate gold(III) complexes adopt similar square-planar geometries as cisplatin; therefore, it was worthwhile to consider that gold(III) complexes might show good anticancer activity. Besides, it was known that gold complexes target proteins unlike cisplatin. Some recent studies have indicated proteasome as the primary target of many gold(III) anticancer complexes [20]. Auranofin (Ridaura); a gold(I) phosphine complex is an orally administered drug (1) which has been used for the treatment of rheumatoid arthritis from a long time ago [21]. It was in the early 1970s and 1980s that this compound and other gold phosphine complexes were found to be active against the growth of cultured tumor cells in vitro [22, 23]. Auranofin does not interact with DNA directly; however, it is known to inhibit DNA, RNA, and protein

252  Advances in Metallodrugs O O

O O O

O O

O 1

O

S

Au

Cl P

Au P 2

Figure 8.3  Chemical structures of some prominent anticancer Au complexes.

synthesis at cytotoxic concentrations. Exposure of cells to auranofin causes several morphological changes like surface membrane changes or cell rounding [24]. Mainly, the complexes containing a phosphine ligand were found to show in vitro cytotoxicity among a series of gold(I) complexes. It was further proved that the replacement or removal of the acetyl groups in the thiosugar moiety of auranofin or a change to other thio sugars did not significantly influence the potency in in vivo antitumor study in mice with ip P388 leukemia [25]. Chloro(triethylphosphine)gold(I), an analog of auranofin (2), in which the thiosugar is replaced by a chlorine atom also showed potent cytotoxicity. Gold(III) complexes have also attracted attention as potential antitumor drugs. Several compounds of this class have displayed strong in vitro antiproliferative effects. The mechanism of action of this class of compounds appears to be the significant inhibition of thioredoxin reductase enzymatic activity [26–28]. A large number of past studies have shown that gold complexes are very attractive as anticancer agents. The chemical structures of 1 and 2 are shown in Figure 8.3.

8.3.1 Complexes with Nitrogen Donar Ligands Gouvea et al. [29] recently documented the cytotoxic activity of three gold(III)  complexes  (3–5) of the fluoroquinolone antimicrobial  agents; norfloxacin, levofloxacin, and sparfloxacin against the A20 (murine lymphoma), B16-F10 (murine melanoma), and K562 (human myeloid leukemia) tumor cell lines as well as the L919 (murine lung fibroblasts) and MCR-5 (human lung fibroblasts) normal cell lines. It was observed that the complexes were more active than their corresponding free ligands. Besides, the complexes displayed moderate binding potential to bovine serum albumin (BSA). The complexes bound to Ct-DNA are more efficiently than their free ligands, which was in well agreement with their cytotoxicity data. Wang and co-workers [30] documented the cellular pharmacological properties of  gold(III) porphyrin (6). The cytotoxicity of  6 was independent

Gold Complexes as Anticancer Agents  253 of its photosensitizing activity. Serum proteins displayed little effects on its activity. Besides, the complex bound to DNA non-covalently quite differently from cisplatin. In addition, inhibition of cell growth was partly through interrupting cell cycle at G0-G1 phase. Coetzee et al. [31] reported the antitumor activities of a series of charged and neutral gold(I) complexes (7–13) of ylideneamine functionalized heterocyclic ligands against cervical carcinoma cells (HeLa). The reported complexes were indeed better cytotoxic agents as compared to their ligands. However, all the complexes were less cytotoxic as compared to cisplatin. Casini and co-workers [32] explored the cytotoxicities of square planar gold(III) complexes that contain functionalized bipyridine ligands. The prepared complexes  showed moderate to good cytotoxicity in vitro towards the A2780 human ovarian carcinoma cell line. Tu et al. [33] reported few gold complexes and tested their cytotoxic effects in vitro to human colon cancer and concluded that one of the complexes may act as new potential therapeutic drug for colon cancer. In another study, Patel et al. [34] synthesized few square planer Au(III) complexes of type [Au(A(n))Clx]·Cly and their in vitro cytotoxic studies. The results revealed significant cytotoxic properties. The chemical structures of 3–13 are shown in Figure 8.4.

HN

Cl Au Cl

CH3

N

H3C Cl

N

N

O

3

Cl

N O

4

Cl

HN

Cl Cl Au

H3C H3C

OH

F

O

N N Au N N

CH3

O

N

OH

F

Cl Cl Au

O

CH3 N H N S Au

7

Cl OH

F 5

CH3 H N N N Au

N

N

NH2 O

O

CH3 H N N S Au

H3C

8

9

6 CH3 N N 10

CH3 H

N NO3 Au PPh3

N S 11

H N Au NO3 PPh3

H3C

CH3 N S

H N Au NO3 PPh3 12

CH N

H N Au

N CH3

N

N

NO3

13

Figure 8.4  Chemical structures of anticancer Au complexes with nitrogen donar ligands.

254  Advances in Metallodrugs

8.3.2 Complexes with Sulphur Donar Ligands Lessa et al. [35] reported the cytotoxicity and thioredoxin reductase activity of gold(I) complexes (14–18) of 2-acetylpyridine thiosemicarbazone and its N(4)-methyl and N(4)-phenyl derivatives, as well as N(4)-phenyl2-benzoylpyridine thiosemicarbazone. The complexes were active against Jurkat (immortalized line of T lymphocyte), HL-60 (acute myeloid leukemia), MCF-7 (human breast adenocarcinoma), and HCT-116 (colorectal carcinoma) cancer cell lines. Interestingly, Jurkat and HL-60 cells were more effected as compared to MCF-7 and HCT-116 cells. The complex 18 displayed more activity as compared to auranofin against both leukemia cells. Besides, all the reported complexes induced DNA fragmentation in HL-60 and Jurkat cells indicating their pro-apoptotic potential. Inhibition of thioredoxin reductase (TrxR) was suggested as the possible mechanism of action of 18. Ronconi et al. [36] reported the solution behavior, electrochemical properties and biological activities of several gold(III) dithiocarbamate derivatives (19–22). The dithiocarbamates were fourfold more cytotoxic than cisplatin in vitro. The complexes underwent complete hydrolysis within a period of 1 h. Besides, the complexes showed high reactivity toward some biologically important isolated macromolecules causing inhibition of the synthesis of both DNA and RNA, in addition, to the induction of DNA lesions faster than cisplatin. Out of the above studied complexes, [Au(III) Br2(ESDT)] (AUL12) (22) was reported to display promising antitumor activity in vitro. Besides, this complex overcame both acquired and intrinsic resistance of some tumors towards cisplatin. An extension of this work by Marzano and co-workers [37] reported thatAUL12 had higher anticancer activity than cisplatin toward all the murine tumor models, in vivo with up to 80% inhibition of tumor growth. Moreover, low acute toxicity levels (lethal dose, LD50 = 30 mg kg−1) and reduced nephrotoxicity were observed with the treatment with this complex. These results suggested AUL12 as a suitable candidate for clinical trials. Saggioro et al. [38] demonstrated the potential of the above complexes (19–22) to induce cancer cell death via apoptotic and non-apoptotic routes. The reported complexes inhibited thioredoxin reductase activity in addition to the generation of free radicals, modification of some mitochondrial functions. The authors suggested that the deregulation of the thioredoxin reductase/ thioredoxin redox system as a major mechanism of the anticancer activity of the complexes. Kouodom and co-workers [39] reported gold(III)-­ dithiocarbamate derivatives of oligopeptides as potential anticancer agents. Among the screened complexes, (22) was the most effective towards PC3, 2008, C13, and L540 with IC50 value lower than cisplatin. Besides, it showed

Gold Complexes as Anticancer Agents  255 CH3

N N

Cl

NH

NH2

S

Au

N H

.CH3OH

Cl

Au

14

H3C

NH

S

NH CH3

Cl

C

S S

Au

Cl Cl

H3C

N

N

Au

S

NH Cl

C

S S

Br

Au

Br

S Br H2C N C Au Br CH3 S 21

Au Cl

S S

C N

Au

S

NH NH

H2 H3C C

O S Cl H2C N C Au CH3 S Cl 20

O H3C CH3 Cl

N

17

19 O

N H

NH

16

H3C

18

H2 H3C C

N

CH3

N H

15

H3C N

CH3

N H

H N

O

OC(CH3)3 O H3C CH3

22

Figure 8.5  Chemical structures of anticancer Au complexes with sulphur donar ligands.

no cross-resistance with cisplatin and inhibited tumor cell proliferation by the induction of late apoptosis/necrosis. The chemical structures of 14–22 are shown in Figure 8.5.

8.3.3 Complexes with Phosphorus Donar Ligands Schuh et al. [40] prepared gold(I) alkynyl complexes (23–25) by reacting propargyl ethers, viz., 7-chloro-(4-propargyloxy)quinoline, 1-propargyloxynaphthalene, and 2-propargyloxy benzophenone with [AuCl(PPh3)]. The complexes showed antiproliferative effects on human cancer cells with IC50 values ranging from 0.4 to 12 µM against CH1 and SK-OV-3 cells (ovarian carcinoma), HeLa cells (cervical carcinoma), and SW480 cells (colon carcinoma) human cancer cell lines. Humphreys and co-workers [41] reported the anticancer activities of gold(I) chloride adducts of 1,3-bis­ (di-2-pyridylphosphino)propane. The complex (26) was selectively toxic to breast (MDA-MB-468) cancer cells. This work was further extended by Rackham et al. [42] who reported that 26 induced apoptosis via the mitochondrial pathway involving mitochondrial membrane potential depolarization, glutathione pool depletion and caspase-3 and caspase-9 activation. Besides, 26 inhibited both thioredoxin and thioredoxin reductase and this effect was more profound in breast cancer cells and this was accounted for the selective cell death seen in the breast cancer cells. Further insights into

256  Advances in Metallodrugs O

Au PPh3

N

Cl

23

Au PPh3

O

24

N N

O

O

N

Au PPh3 N

25

N P P

P Au

N P

N

Cl

N N 26

Figure 8.6  Chemical structures of anticancer Au complexes with phosphorous donar ligands.

the mechanism of action of this complex were provided by Wedlock and co-workers [43]. They reported the subcellular distribution of this complex in situ in human breast cancer cells using nano-scale secondary ion mass spectrometry. It was observed that the subcellular distribution of gold was associated with sulphur-rich regions in the nucleus and cytoplasm, indicating the mechanism of action of Au(I) complexes involves the inhibition of thiol-containing protein families, such as the thioredoxin system. The chemical structures of 23–26 are shown in Figure 8.6.

8.3.4 Complexes with Sulphur-Phosphorus Donar Ligands Bagowski and co-workers [44] documented a series of  gold(I) phosphine  complexes (27-30) bearing a naphthalimide ligand. The complexes exhibited strong antiproliferative effects against MCF-7 and HT-29 cancer cells. Among the tested complexes, 30 displayed solvent-dependent fluorescence emission, uptake into the organelles of tumor cells and anti-­ angiogenic effects in vivo. Barreiro et al. [45] reported the cytotoxic properties of Triphenylphosphine gold(I) sulfanylpropenoates (31–40) towards Hela-229, A2780, cisplatin-resistant mutant A2780cis and ovarian carcinoma cells. All the complexes showed the maximum activity against HeLa cells with IC50 values in the low micromolar range (3.3–12.0 µM). The best IC50 value (3.3 µM) was obtained for 32. The same group tried to explore the anticancer properties of more similar types of complexes [46], like in vitro antitumor activities of diisopropylammonium and triethylammonium salts of triphenylphosphinegold(I) sulfanylcarboxylates (41–50) against HeLa229, A2780, and A2780cis cell lines. The diisopropylammonium derivatives

Gold Complexes as Anticancer Agents  257 were more active, in particular against the A2780 cis cell line and displayed a high potential to circumvent the cellular cisplatin resistance. Vergara et al. [47] reported the in vitro cytotoxicity and thioredoxin reductase inhibition of a series of gold(I) complexes (51–57) containing hydrophilic phosphine ligands against human ovarian cancer cell lines A2780 both cisplatin sensitive or resistant. All the complexes displayed promising antiproliferative effects, against the cisplatin-resistant cell line with IC50 in 4–16 mm range. Besides, the compounds inhibited cytosolic and mitochondrial thioredoxin reductases in vitro. It is indeed that the central functions of the thioredoxin system make it an attractive target for anticancer drug development. The chemical structures of 27–57 are shown in Figures 8.7–8.7.3. H3C S Au P CH3 H3C

O

N

N O

N O

28

O

N

(H3C)3C

CH2 CH 3 S Au P CH2 H2C CH3

N O

(H3C)3C S Au P C(CH3)3

O

N

N

27

H3C

O

S Au P

N O

29

30

Figure 8.7  Chemical structures of anticancer Au complexes with sulphur-phosphorus donar ligands. COOH C H

S Au

31

PPh3

O

N

C H 32

C

S Au

S Au

PPh3 35

S Au

C C H

PPh3

37

38 Br

33

S Au

OCH3 C H 36

PPh3

PPh3

COOH C S Au

PPh3 COOH

COOH

C S Au

S

C

OH

COOH C H

PPh3

C H

COOH C C H

34

H3CO

C

Cl

COOH C H

COOH

COOH

C

S Au

HO PPh3

C H

C S

Au

39 PPh3

OH COOH C H

Br

40

C S Au

PPh3

Figure 8.7.1  Chemical structures of anticancer Au complexes with sulphur-phosphorus donar ligands.

258  Advances in Metallodrugs O

O

O

O

(CH2)2CHNH2+ Ph3P Au S

(CH2)2CHNH2+ Ph3P Au S O

41 O

S

42 O

O

O

(CH2)2CHNH2+ Ph3P Au S

(CH2)2CHNH2+ Ph3P Au S

43

44

N O

O O

45

46

Cl

O

(CH2)2CHNH2 Ph3P Au S 47

Br

HO

Br

O

O +

O

(CH2)2CHNH2+ Ph3P Au S

(CH2)2CHNH2+ Ph3P Au S

O

(CH2)2CHNH2+ Ph3P Au S

OH

48 HO O

O O

O

(CH2)2CHNH2+ Ph3P Au S

(CH2)2CHNH2 Ph3P Au S +

49

OCH3

50 H3CO

Figure 8.7.2  Chemical structures of anticancer Au complexes with sulphur-phosphorus donar ligands.

N

S Au P

N N

N

N

N

51

H3C

CH3 N S Au P N N

N 55

N

O N N

52 O

N

S Au P

N

H3C

O

NaO3S N 56

S Au P 53

H3C

SO3Na S Au P

SO3Na

CH3 N N

N

N H3C

O

S Au P

N

S 57

N

N

54

NaO3S N

N

SO3Na S Au P

SO3Na

Figure 8.7.3  Chemical structures of anticancer Au complexes with sulphur-phosphorus donar.

Gold Complexes as Anticancer Agents  259

8.3.5 Organometallic Gold Complexes Schuh and co-workers [48] reported the antiproliferative properties of a series of gold(I)  complexes  with 1,3-substituted imidazole-2-ylidene and benzimidazole-2-ylidene ligands on human ovarian cancer cells, sensitive, and resistant to cisplatin (A2780S/R) as well as on the nontumorigenic human embryonic kidney cell line (HEK-293T). Good antiproliferative activities and potent selective TrxR inhibition properties of all the complexes have been observed. The most effective TrxR inhibitors were observed to induce extensive oxidation of thioredoxins. Rubbiani et al. [49] reported the antitumor properties and thioredoxin reductase inhibitory activities of gold(I) complexes (58–61) with benzimidazole derived N-heterocyclic carbene ligands. Interestingly, the complexes inhibited TrxR selectively as compared to glutathione reductase. Besides, complexes displayed significant antiproliferative activities in cultured tumor cells. In another work, Liu et al. [50] documented a series of neutral NHC gold halide complexes. The pharmacological studies indicated high growth inhibitory effects of the complexes on MCF-7, MDA-MB 231, and HT-29 colon cancer cell lines, which were dependent on the presence of C4, C5-standing aromatic rings. However, the presence of methoxy groups on these rings had no effect on the inhibitory activities, whereas fluorine substituents on the ortho-positions were contributing positively to the activity on MCF-7 and MDA-MB 231 cells. Out of the screened compounds, the most active compound ligands 62 {bromo[1,3-diethyl-4,5-bis(2-fluorophenyl)-1,3-dihydro2H-imidazol-2-ylidene]gold(I)} exhibited greater activity as compared to cisplatin. Various biochemical analyses excluded thioredoxin reductase inhibition, DNA interaction and growth inhibition and inactivation of COX enzymes as the biological targets of these complexes. In another study, Yan et al. [51] synthesized a panel of stable [Au(R-C-N--C)(N-heterocyclic carbene)](+) complexes and revealed their prominent in vitro anticancer properties. The complex [Au(C--N--C)(IMe)] CF(3)SO(3) (1, IMe = 1,3-dimethylimidazol-2-ylidene) significantly poisons topoisomerase I in vitro and suppresses tumor growth in nude mice model. Hickey and co-workers [52] designed a family of lipophilic gold(I) N-heterocyclic carbene complexes as toxic agents to cancer cells via mitochondria targeted pathway. The chemical structures of 58–62 are shown in Figure 8.8.

260  Advances in Metallodrugs

CH3 N Au Cl N CH3 58

H3C

CH2 N Au Cl N H3C CH2 59

CH2 N Au Cl N CH2 60

CH N N CH 61

Au Cl

F H 2 H3C C N

F N

F

Au Br 62

Figure 8.8  Chemical structures of organometallic anticancer Au complexes.

8.3.6 Miscellaneous Maiore et al. [53] reported the solution behavior and cytotoxicity of a series of gold(III) complexes (63–66) of 2-substituted pyridines. The reported complexes simply behave as classical prodrugs wherein the activation of the metal center occurs by the release of the labile chlorido ligands without altering the rest of the molecule. Besides, the reported compounds reacted with the model protein cyt. c leading to extensive protein metalation. The compounds exhibited remarkable cytotoxic activities against the human ovarian carcinoma cell lines; A2780/S and A2780/R with IC50 values ranging from 1.43 to 6.18 μM in the sensitive cell line and from 1.59 to 10.86 μM in the resistant one, respectively. More importantly, all the reported compounds had the ability to overcome cisplatin resistance. In order to understand the mechanism of action of gold-based coordination complexes, Vela et al. [54] tried to explore cellular events triggered by three different iminophosphorane-organo gold(III) complexes (67–69) in leukemia cells. It was observed that the three investigated complexes displayed higher toxicity against leukemia cells when compared to normal T-lymphocytes. In addition, necrosis and apoptosis was induced by both complexes 67 and 68, while the complex 69 was mainly apoptotic. Moreover, production of reactive oxygen species (ROS) at the mitochondrial level was thought of as a critical process in the antitumor effect of these complexes. Wang and co-workers [55] reported the cytotoxicities of a series of eight new gold(III) complexes (70–76) of 5-aryl-3-(pyridin-2-yl)-4,5-dihydropyrazole-1-carbothioamide derivatives. The complexes 70–76 exerted cytotoxic effects against HeLa and A549 cell lines among which the  complexes 71–76 displayed higher cytotoxicity (IC50 range = 2.55–5.14) than cisplatin (IC50 = 6.86) against HeLa cell line. Cytotoxic gold compounds hold today great promise as new pharmacological agents for treatment of human ovarian carcinoma; yet, their mode of action is still largely unknown. To shed light on the underlying molecular mechanisms, Guidi et al. [56] performed 2D-DIGE analysis to identify differential protein expression in a cisplatinsensitive human ovarian cancer cell line (A2780/S) following treatment with two

Gold Complexes as Anticancer Agents  261 representative gold(iii) complexes that are known to be potent antiproliferative agents, namely, AuL12 and Au(2)Phen. Software analysis using DeCyder was performed and few differentially expressed protein spots were visualized between the three examined settings after 24 h exposure to the cytotoxic compounds, implying that cellular damage at least during the early phases of exposure is quite limited and selective, reflecting the attempts of the cell to repair damage and to survive the insult. The potential of novel proteomic methods to disclose mechanistic details of cytotoxic metallodrugs is herein further highlighted. Different patterns of proteomic changes were highlighted for the two metallodrugs with only a few perturbed protein spots in common. Using MALDI-TOF MS and ESI-Ion trap MS/MS, several differentially expressed proteins were identified. Two of these were validated by western blotting: Ubiquilin-1, responsible for inhibiting degradation of proteins such as p53 and NAP1L1, a candidate marker identified in primary tumors. Ubiquilin-1 resulted over-expressed following both treatments and NAP1L1 was downexpressed in AuL12-treated cells in comparison with control and with Au(2) Phen-treated cells. In conclusion, we performed a comprehensive analysis of proteins regulated by AuL12 and Au(2)Phen, providing a useful insight into their mechanisms of action. The chemical structures of 63–76 are shown in Figure 8.9. O

O Au

Cl

N

N

N

Au

Cl

Cl 63

S Au S

N H3C

67

Cl Au

Cl Au

S

74

PF6- PR3

N

PPh2 N Ph PF6-

Au Cl

3.193

Cl Au

S 71

NH

NO2

N N

N Cl Au

Cl

N NH

Cl Au

N

N

NH

O

N Cl Au

N

S

72

Cl Au

N

N N S NH 75

CH3

N

S

NH 69

N

NH

N

N

N N Cl Au S NH 76

Figure 8.9  Chemical structures of other anticancer Au complexes.

O

N

S

NH 73

O N

Cl

R={Cp(m-C6H4-SO3Na)2}

N

S 70 N

PPh2 N Ph

68

Cl N

Cl

64

PPh2 Cl Au N Ph H3C Cl

N

O N O CH3 N Cl N Au CH3 Cl Au N CHMe2 Cl CHMe2 O OH 65 66

N

262  Advances in Metallodrugs

8.4 Nano-Formulations of Gold Complexes Nano-sized drug delivery systems are being increasingly demanded in pharmaceutical industry. These nano-formulations remarkably improve the pharmacokinetics and biodistribution of drugs leading to reduction of side effects and improvement of patient compliance. Since the approval of liposome containing doxorubicin in 1995, a large number of nano-­ formulations, such as polymer-drug conjugates, dendrimers, or inorganic nanoparticles, have entered clinical trials [57]. Pharmaceutical formulations including nanosized drugs are, generally, referred to as nanopharmaceuticals and are quite significantly beneficent to the patient in comparison to the conventional drugs. Nano-formulations have several advantages such as enhanced solubility, oral bioavailability, dose proportionality, and reduced food effects and suitability for administration via all routes [58]. The antitumor activity of organo-gold compounds is a focus of research from the past two decades. A variety of  gold  stabilizing ligands such as vitamins and xanthanes have been prepared and explored for their “chelating effect” as well as for their antitumor activity. Dithiocarbamates (DTC’s) and their metallic conjugates have been well explored for their antiproliferative activities. Yan et al. [59] reported encapsulation of gold(III) complexes with porphyrin and confirmed their rapid drug release properties with improved anticancer efficacy. Hosta and co-workers [60] in 2009 reported the enhanced antitumor activity of drug Kahalaide F which was conjugated with gold nanoparticles. In another work, reported by Ahmed et al. [61], glycopolymer conjugates based on DTC’s were prepared by reversible addition-fragmentation chain transfer polymerization and then modified with gold(I) phosphine. The authors tested the conjugates for in vitro toxicity in both normal and cancer cell lines. While as, the Au(I) phosphine conjugated cationic glycopolymers of 10 kDa and 30 kDa were evaluated for their cytotoxicity profiles using MTT assay. Overall, it was concluded that these polymers based on DTC’s and their gold conjugates indeed show higher accumulation as well as higher cytotoxicity to cancer cells under hypoxic conditions in comparison to the normoxic ones. Furthermore, it was noticed that hypoxic MCF-7 cells showed significant sensitivity toward the low molecular weight glycopolymer-Au(I)  complexes. Nasrolahi and co-workers [62] synthesized few cyclic peptides which were evaluated as simultaneous reducing and capping agents for generation of cyclic peptidecapped GNP’s (CP-AuNPs). Among them, direct dissolution of cyclic peptides containing alternate arginine and tryptophan [WR](n) (n = 3–5) into an aqueous solution of AuCl(4)(−) led to the formation of CP-AuNPs,

Gold Complexes as Anticancer Agents  263 through the reducing activity of tryptophan residues and attraction of positively charged arginine residues towards chloroaurate anions in the reaction environment. Additionally, flow cytometry assessment showed that in the presence of [WR](4)-AuNPs, the cellular uptake of fluorescence labeled stavudine, emtricitabine, and lamivudine was significantly enhanced in human ovarian adenocarcinoma (SK-OV-3) cells. Furthermore, confocal microscopy nailed out that the presence of the [WR](4)-AuNPs enhanced the retention and nuclear localization of doxorubicin in SK-OV-3 cells after 24 h. Overall, the data suggested that gold complexes  can be used as potential non-covalent prodrugs for delivery of antiviral and anticancer agents. Pearson et al. [63] reported the increased anti-proliferative properties of a  gold  complex with a sugar ligand against OVCAR-3 (human ovarian carcinoma cells). The nano form of gold conjugates was prepared by post-modifying RAFT glycopolymers. Recently, Kouodom et al. [64] designed few gold(III) complexes on purpose to obtain anticancer candidates based on peptides. The candidates target two peptide transporters (namely, PEPT-1 and PEPT-2) upregulated in several tumor cells. According to in vitro cytotoxicity studies, few complexes turned out to be the most effective toward all the human tumor cell lines evaluated (PC3, DU145, 2008, C13, and L540), with lower IC50 values than cisplatin. More importantly, they showed no cross-resistance with cisplatin itself and were proved to inhibit tumor cell proliferation by inducing either apoptosis or late apoptosis/necrosis depending on the cell. Of course, nano-formulations of gold complexes are improved candidates for the chemotherapy of cancer. Obviously, research should be encouraged in this direction to develop a nano-sized gold complexes as future anticancer drugs.

8.5 Future Challenges and Perspectives Last few decades have seen extensive scientific revolution in almost every sphere of science and technology. Despite of this scientific revolution, the treatment of cancer still remains a major challenge [65]. Of course, cancer is a huge threat to human beings globally and a serious challenge to our society. In the present scenario, we are not fully developed for eradicating cancer. Moreover, the anticancer drugs available in the market are unable to cure cancer; especially in its late stages and as a result, higher death cases as compared to the survival cases are reported. Besides, the long-term uses of the available anticancer drugs induce resistance in cancer cells that greatly limits their application. The costs of the market selling anticancer

264  Advances in Metallodrugs drugs are too high and hence cannot be afforded by the common people. The way that could lead to the development of novel metal anticancer drugs has many obstacles. The main obstacle seems to be that no general guideline towards the synthesis of new active metal complexes has been established, since the discovery of active complexes was accidental. In many cases, a few structure-activity relationships were found; however, there are no established general rules. Therefore, which other metal complexes are expected to be active and should be investigated in near future still remains a mystery. Seemingly significant advances have been made for understanding the molecular etiology of cancer, but ideal therapeutic strategies against this disease are still missing. As a consequence of these facts, it becomes very crucial to expedite the process of the development of new therapeutic agents against cancer [66]. Developing new and efficient drugs for the treatment of diseases has been the prime goal for different areas of research including natural products chemistry, molecular biology and biochemistry, pharmacology, and medicinal chemistry [67]. Literature suggests that the therapeutic potential of metal complexes can be harnessed for the design of novel and efficient anticancer agents. Chemical and biological properties of metal complexes are quite different from those of purely organic molecules. Metal ions from the first, second, and third row transition series are quite preferred because of their variable oxidation states and coordination numbers and the potential to coordinate with a wide variety of ligands. Cisplatin and its analogs (carboplatin and oxaliplatin) have proved the fact that metal complexes can play important roles in cancer treatment regimes. So, it is the time to explore other transition metal complexes as anticancer drugs. Targeting and activation strategies should be encouraged for the development of future generations of drugs which can overcome some of the disadvantages associated with cisplatin therapy. The drugs obtained should be bestowed with no or reduced side effects, broad spectrum of activity, and should be capable to avoid the occurrence of drug resistance [68]. It is very important to understand the parameters by which ligands control the reactivity of transition metal ions, and also the reciprocal effects which metal ions can have on the properties of ligands since both these identities play important roles in the recognition of target sites. Some of the modern theoretical methods such as Density Functional Theory and techniques like high resolution electrospray mass spectrometry, multinuclear polarization transfer NMR spectroscopy can improve our understanding of the chemical and biochemical reactivity of metal complexes and the construction of meaningful structureactivity relationships. Thus, studies of the chemistry of metal complexes under physiologically-relevant conditions (e.g. biological screening

Gold Complexes as Anticancer Agents  265 conditions) become very important. Metal complexes with targeted action on cancerous cells and considerably lower side effects can be obtained by synthesizing their nano identities. Trapping complex ions into nano cages may avoid their poor bioavailability with selective, specific and fast action increasing the longevity of patients [69]. Nano identities of gold as well as other metal-based drugs may be expected to cure a number of cancers with fewer side effects which is the need of future [70]. A few reports are in favor of the fact that synergistic effects in several cancer cases have been observed by the use of metal complexes in combination with other agents. It is therefore expected that combination therapies may be tried to get novel drug combinations in near future. Presently, we have got software and simulation programmes, which allow us to estimate drug therapeutic efficiency even before their synthesis. Therefore, the synthesis of such computer simulated drugs having good bioavailability, maximum solubility, remarkably less side effects, and higher efficiency need to be designed and developed. Drug delivery systems are known to work as innovative vehicles for the transport and targeting of anticancer drugs. Metallodrugs should be delivered to target sites by means of nanodrug delivery systems decorated with cell specific antibodies. Generally, multiple complex biochemical pathways are implicated in diseases like cancer whose successful treatment usually depends on pharmaceutical intervention at multiple pathways, and, often with a combination of different drugs. Designed multiple ligands (DMLs) that act at multiple biological targets may be quite helpful in the eradication of the deadly disease cancer [71]. Therefore, it might be suggested that the development of metallodrug-based DMLs might be effective in the fight against cancer. Keeping these points in view, there is an urgent need to understand the molecular mechanism of apoptosis induced by new metallodrugs and to synthesize their nano counterparts and see how it works. Scientists and oncologists all over the world should work together to develop hundred percent safe metallodrugs (either in nano regime or the normal size range) for all sorts of cancers. Briefly, the overall observation, experience, and literature update dictates that the future of metallo­drugs is quite bright in the treatment of cancer. Let’s hope for the best future of metallodrugs for the service of mankind in the coming time.

8.6 Conclusion Really, the discovery of the anticancer properties of cisplatin stimulated the development of metal-based drugs for the treatment of cancer. Several ­metal-based drugs were developed with promising anticancer properties and

266  Advances in Metallodrugs many of them are being sold in market in the present time. Irrespective of this grand achievement, we do not have a drug that can cure cancer at its late stages without serious side effects. This idea of the development of an ideal anticancer drug tempted scientists all over the world to explore the potential of metal complexes as anticancer agents. Au complexes have been very central in this aspect of metal drug development. Despite of the fact that many of the Au drugs are superior to non-Au drugs as candidates for treating cancer, there is an ever increasing demand for the development of metal anticancer drugs. Recently, much research is being carried out all over the world on the development of metal complexes for the treatment of cancer. Cancer being a deadly fatal disease affecting the social and economic status of such patients drastically needs to be eradicated as soon as possible. Definitely, the future of Au anticancer drugs is quite bright since this is a step in the right direction towards the eradication of the deadly havoc of cancer.

Acknowledgements The authors are highly grateful to Department of Biotechnology, Central University of Kashmir and Department of Chemistry, National Institute of Technology, Jammu and Kashmir, India, for providing good research facilities and cordial academic environment.

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9 Recent Developments in Small Molecular HIV-1 and Hepatitis B Virus RNase H Inhibitors Fenju Wei1, Dongwei Kang1, Luis Menéndez-Arias2*, Xinyong Liu1† and Peng Zhan1‡ Department of Medicinal Chemistry, Key Laboratory of Chemical Biology, Ministry of Education, School of Pharmaceutical Sciences, Shandong University, Jinan, China 2 Centro de Biología Molecular “Severo Ochoa” (Consejo Superior de Investigaciones Científicas & Universidad Autónoma de Madrid), Madrid, Spain 1

Abstract

Finding new multifunctional metal-chelating inhibitors to be potentially used in therapy is of utmost importance in drug development. Mechanistic studies have shown the remarkable antiviral potency of compounds that exert their inhibitory activity through chelating metal cations found in the active sites of enzymes required for virus propagation. This chapter focuses on strategies applied to the discovery of drugs chelating Mg2+ cations in the ribonuclease H (RNase H) active site, as a relevant target to block human immunodeficiency virus (HIV) and hepatitis B virus (HBV) replication. We summarize recent developments in the field, while presenting the chemical structures, structure-activity relationships, and binding modes of novel RNase H inhibitors. Keywords:  Ribonuclease H, reverse transcriptase, HIV, hepatitis B virus, integrase, active site inhibitors

9.1 Introduction Human immunodeficiency viruses type 1 and type 2 (HIV-1 and HIV-2, respectively) and hepatitis B viruses (HBV) are important pathogens that *Corresponding author: [email protected] † Corresponding author: [email protected] ‡ Corresponding author: [email protected] Shahid-ul-Islam, Athar Adil Hashmi and Salman Ahmad Khan (eds.) Advances in Metallodrugs: Preparation and Applications in Medicinal Chemistry, (273–292) © 2020 Scrivener Publishing LLC

273

274  Advances in Metallodrugs use reverse transcription to replicate their genomes [1, 2]. According to the most recent worldwide estimates, there are about 38 million people living with HIV (www.unaids.org), while more than 250 million people are chronically infected with the HBV. In 2018, HIV/AIDS was globally responsible for 770,000 deaths, and it has been estimated that 20% to 30% of the adults that are chronically infected with HBV develop cirrhosis and liver cancer [3]. Despite recent advances in antiviral therapy, the currently available treatments should not be considered fully satisfactory due to their high cost, severe side effects, and the development of viral drug resistance [4–6]. In this scenario, novel drugs are necessary to improve current therapies and reduce the global burden of infectious disease, including HIV/ AIDS and HBV infection.

9.1.1 Activity and Function of HIV and HBV RNases H Ribonucleases H (RNases H) are endonucleolytic enzymes that catalyze the cleavage of RNA in RNA/DNA substrates. They are evolutionarily related to retroviral integrases, DNA transposases, resolvases, and many nucleases [7, 8] and play important roles in the replication of prokaryotic and eukaryotic genomes [9], as well as in the replication of reverse-transcribing viruses [2]. From a mechanistic point of view, there are important differences in the process of reverse transcription in HBV and HIV [2]. However, the RNase H activity of the viral polymerase is required in both cases to degrade viral genomic or pregenomic RNA as a necessary step to facilitate the formation of double-stranded DNA. In addition, during reverse transcription, the RNase H plays a role in template switching [10], a property of retroviral and hepadnaviral reverse transcriptases (RTs) that facilitates the formation of long terminal repeats (LTRs) in retroviruses, and the circularization of the HBV DNA genome [2].

9.1.2 The Metal-Chelating RNase H Active Site The HIV RNase H active site shares with prokaryotic and eukaryotic RNases H a highly conserved “DEDD” motif that in the case of HIV-1 contains the catalytic carboxylate residues Asp443 (D443), Glu478 (E478), Asp498 (D498), and Asp549 (D549) (Figure 9.1). These amino acids coordinate two divalent cations required as cofactors to hydrolyze the RNA substrate. The substitution of any of the four highly conserved residues leads to the loss of RNase H activity [11–13]. Most HIV RNase H inhibitors coordinate the two catalytic Mg2+ ions in the RNase H active site, usually through a triad of Lewis basic atoms or electron donors.

Molecular HIV-1 and HBV RNase H Inhibitors  275 Base Base OH O

O O

OH O O

O

O

O

P

O

O

O

Mg2+

E478

O

P

OH Mg2+ O

O O

H

D498

O 2

O

O

O

D549

D443

O

Figure 9.1  The two-metal ion mechanism of catalysis for HIV-1 RNase H activity [14].

The RNase H domain is located at the C-terminal region of the HIV RT and the HBV polymerase. However, HBV and HIV RNases H show limited homology with only 23% amino acid sequence identity [15]. The HBV RNase H has been successfully expressed as a fusion recombinant protein containing an N-terminal maltose-binding protein (MBP) and a C-terminal hexahistidine (H6) tag [16]. Catalytic residues (Asp702, Glu731, Asp750, and Asp790) equivalent to those indicated above for HIV-1 RNase H have been identified by introducing non-conservative substitutions in the DEDD motif (Figure 9.2) [17–19]. The role of the HBV RNase H is to degrade the pregenomic viral RNA after being transcribed into DNA by the viral polymerase in order to facilitate synthesis of the positive strand of DNA. Experimental evidence indicates that RNase H inhibition has effects on the synthesis of both viral DNA strands (i.e., plus and minus polarity) [20]. The HBV RNase H is Mg2+-dependent, and all known HBV RNase H inhibitors have metal-coordinating moieties that could chelate the active site cations. Likewise, all HBV Polymerase 688aa TP

Sp

157aa RT

RNase H

Recombinant RNase H MBP

D E D

D

H6

Figure 9.2  The HBV RNase H. TP, terminal protein domain that primes DNA synthesis; Sp, spacer domain; RT, reverse transcriptasve domain; RNase H, RNase H domain; MBP, maltose-binding protein; H6, hexahistidine tail.

276  Advances in Metallodrugs HBV RNase H inhibitors identified to date feature a three-oxygen pharmacophore or an equivalent structure as a triad that binds the Mg2+ cations required for catalysis. Disrupting the geometry of the active site residues by introducing chelators or altering interactions of the amino acid side chains of the “DEDD” motif is considered a valid strategy to inhibit HBV and HIV RNases H [21], converting these enzymes into attractive targets for the development of antiviral drugs.

9.2 RNase H Inhibitors and Strategies in the Discovery of Active Compounds 9.2.1 High-Throughput Screening High-throughput screening (HTS) of large collections of molecules has been a valuable technique in the search for active compounds and remains as a major procedure towards the identification of novel hits in drug discovery [22]. Furthermore, this approach has demonstrated its efficiency and flexibility in the identification of valuable lead compounds. The dihydroxycoumarin F3284-8495 (1) was identified by Himmel et al. by HTS. F3284-8495 showed low micromolar HIV-1 RT RNase H inhibitory activity (IC50 = 4.8 μM at pH 7.4), but was unable to block DNA synthesis catalyzed by the RT at 10 μM concentration [23]. After further analysis of the co-crystal structure of 1 bound to the RNase H active site, researchers replaced the ethanoic acid moiety by a series of larger substituents, in order to introduce additional interactions within the RNase H domain and improve their inhibitory activity. A series of analogs of compound 1 was obtained [23]. As expected, compounds F3385-2581 (2) and F3385-2590 (3) (Figure 9.3), containing a piperazine ring linked to one or multiple aromatic rings instead of the ethanoic acid substituent, showed inhibitory activity against an engineered HIV RNase H domain known as

HO HO

O

O

HO O HO

OH F3284-8495 (1)

O

O

HO N

N

F3385-2581 (2)

O

HO

O Cl N

N

F3385-2590 (3)

O

OH

HO β-thujaplicinol (4)

HIV RNase H inhibitors

Figure 9.3  Structures of HIV RNase H inhibitors dihydroxycoumarins (1–3) and β-thujaplicinol (4).

Molecular HIV-1 and HBV RNase H Inhibitors  277 p15-EC RNase H at submicromolar levels (IC50 = 0.1 μM and 0.2 μM for compounds 2 and 3, respectively). Apart from the inhibitors described above, several reports have demonstrated the HIV-1 RNase H inhibitory activity of tropolone derivatives [24–26]. Among them, a tropolone derivative with a 7-OH substitution discovered in a HTS campaign using National Cancer Institute libraries of pure natural products was found to inhibit retroviral, bacterial, and human RNases H [26]. The compound, known as β-thujaplicinol (4), is found in the heartwood of western redcedar, a coniferous tree of the Cupressaceae family, and showed an IC50 of 0.21 μM in HIV-1 RNase H inhibition assays (Figure 9.3) [24]. Later studies showed that β-thujaplicinol was also an inhibitor of the HBV RNase H. Tavis et al. demonstrated its antiviral activity using a cellbased phenotypic screening assay. Major stabilizing interactions of the HBV RNase H-inhibitor complexes are probably electrostatic and appear to be primarily established between the chelating moiety of the ligands and the polar catalytic center of the enzyme (Figure 9.4) [19]. Preliminary structure-activity relationships (SAR) using hydroxytropolones indicated that the intact α-tropolone moiety is needed because deleting one of the three oxygens on the tropolone ring ablates inhibition. This implies that hydroxytropolones inhibit the HBV RNase H by the metal-chelating mechanism employed against the HIV RNase H. The most active hydroxytropolone was the least-substituted molecule, compound 5, which has a single isopropyl group with an IC50 value of 5.9 μM against HBV [27]. The substituents at positions R1, R2, and R3 should be short for O HO

O

OH

O 6 IC50 = 34.6 µM

5 IC50 = 5.9 µM HO

OH

HO

O

O

OH O

OH

HO

Metal chelating moiety

HO

HO 8 IC50 > 100 µM

Mg2+ O

α-hydroxutropolone moiety is needed O

O O

O 7 IC50 = 29.6 µM

Mg2+ O O +

R3 R2 R1

Preliminary SAR

≥ 1 small substituents are needed

HBV RNase H inhibitors

Figure 9.4  Structures of HBV RNase H inhibitors (5–8) and their metal chelating moiety and preliminary SAR.

278  Advances in Metallodrugs substantial activity (length equal or smaller than four atoms) [28]. These size limitations suggest that the HBV RNase H active site is probably narrower than the HIV RNase H active site. A narrower active site is expected to facilitate the design of compounds with adequate specificity for the RNase H [15].

9.2.2 Design Based on Pharmacophore Models A distinct pharmacophore featuring a highly privileged biaryl moiety connected to the chelating core by a one atom linker seems to be common to many RNase H inhibitors and could be the key for tight binding to the RNase H active site. Based on this pharmacophore model for RNase H inhibition, the 3-hydroxypyrimidine-2,4-dione (HPD) chemotype (Figure  9.5) was redesigned to obtain selective RNase H inhibitors [29]. The biaryl group with different substituents was introduced at the C6 position of the HPD using different linkers. Meanwhile, structural simplification was performed, namely, by removing the crucial C5 isopropyl group and substituting the N-1 position with a methyl group (9 and 10) or a hydrogen atom (11). The hydroxypyridonecarboxylic acid pharmacophore, found in the influenza endonuclease inhibitor 12 and in the approved HIV integrase (IN) strand transfer inhibitor dolutegravir 13 (Figure 9.6), has been exploited as a chelating core to design new selective RNase H inhibitors [30]. In 2016, Kankanala et al. reported novel derivatives of the

HO 3 N O

O 5

R

1N 6

O

O

HO O

Structural simplification

R'

O N N H Me

9 IC50 = 0.80 µM

O N N R

O OMe

N

HO

HO O

X

Ar

Ar'

O OMe HO N

N N O Me

10 IC50 = 2.0 µM

HIV RNase H inhibitors

Figure 9.5  Structures of HIV RNase H inhibitors 9–11.

O

OMe N H

N H 11 IC50 = 0.15 µM

Molecular HIV-1 and HBV RNase H Inhibitors  279 OH O O N

OH

N OH O

N 12 Influenza endonuclease inhibitor

O

OH

ASP-443

N

ASP-549 LYS-540

GLU-478 HIS-539

O

OH

N O

N H

O

H N

F

F

14 HIV RNase H inhibitor

O

O S NH 2

ASP-498

Binding pattern

13 O HIV IN inhibitor

Figure 9.6  Structures of compounds 12–14 and the binding mode of compound 14 in the HIV-1 RNase H active site (PDB code: 5J1E).

hydroxypyridonecarboxylic acid scaffold with critical biarylmethyl or N-1 benzyl moieties [31]. Thus, compound 14 (Figure 9.6) was identified as the most effective HIV-1 RNase H inhibitor (IC50 = 0.65 ± 0.05 µM). The recently reported X-ray structure of the HIV-1 RNase H domain bound to 14 represents a significant advance for understanding HIV RNase H inhibition. In the obtained crystal structure, compound 14 chelates two Mg2+ ions through the carbonyl, hydroxyl, and carboxylate groups of the pyrimidone.

9.2.3 Novel Inhibitors Obtained by Using “Click Chemistry” Click chemistry is a term that was first proposed and described by K. Barry Sharpless and colleagues [32]. Cornerstones of click chemistry are high yield, wide scope, less cytotoxic byproducts, high stereospecificity, and a simple reaction. Click chemistry reactions can occur under physiological conditions and the resulting chemical bonds are irreversible. Therefore, click chemistry is widely used for the modification of biomolecules, such as nucleic acids, lipids, and proteins using different compounds [33]. Recently, a series of 4-[4-(aryl)-1H-1,2,3-triazol-1-yl]benzenesulfonamides were prepared using “click chemistry” reactions and identified as novel HIV-1 RNase H inhibitors through an in-house screening campaign. Among them, three compounds (15–17) showed selective potency against the HIV-1 RNase H at micromolar concentrations (Figure 9.7) [34]. Notably, their structures were different from those of previously reported for divalent metal-chelating RNase H active site inhibitors and were considered as good lead compounds for the development of a new generation

280  Advances in Metallodrugs O N

NN

15 IC50 = 63 ± 7 µM

S

NH2 O

F N N N

F O

F S

NH2 O

F

N N N

F O

S

NH2 O

F F

F

16 IC50 = 26 ± 3 µM

17 IC50 = 6.6 ± 0.5 µM

Figure 9.7  Structures of HIV RNase H inhibitors (15–17).

of anti-HIV agents. Compound 17 was the most potent inhibitor in enzymatic assays showing an IC50 of 6.6 μM.

9.2.4 Dual-Target Inhibitors Against HIV-1 Integrase (IN) and RNase H HIV-1 RNase H and IN are metalloenzymes with important functions in the viral life cycle and both of them are mechanistically related, belonging to the polynucleotidyl transferase superfamily [35]. Retroviral INs catalyze the integration of the viral DNA into the genome of infected host cells by the coordinated action of two enzymatic activities (3’-processing and strand transfer). Diketo acid (DKA) inhibitors are active-site Mg2+-binding inhibitors that were initially identified as inhibitors of the strand transfer activity of HIV-1 IN. As expected from the structural similarity between the catalytic sites of HIV-1 RNase H and IN, compounds bearing DKA moieties were frequently identified as dual inhibitors of both enzymes. Thus, the ester derivative RDS1643 (18) has inhibitory activity against both HIV-1 IN and RNase H in the low micromolar range (Figure 9.8) [36]. In addition, several pyrrolyl DKA derivatives, having a quinolinonyl-or a pyrroyl-based scaffold, such as compound 19, have been reported as RNase H inhibitors with dual activity against HIV-1 IN and RT-associated RNase H [37, 38]. Related to DKA derivatives, compound 20 was obtained after adding a 5-N-benzylcarboxamide group to a 3-hydroxypyrimidine-2,4-dione (HPD) core. The molecule was found to be a strong inhibitor of the HIV-1 IN strand transfer and RT-associated RNase H activities with IC50 values of 21 nM and 29 nM, respectively [39]. Related molecules containing

Molecular HIV-1 and HBV RNase H Inhibitors  281 O

OH

O

OH

O O

N O F

N

O

O

N

F

19

RDS1643 (18)

PHE-190

O HO

O

ASP-128

Cl

N H

N N H

O

F

GLU-221

ASP-185

N H 20

Binding mode Dual inhibitors of HIV RNase H and IN

Figure 9.8  Chemical structures of dual inhibitors (18–20) of HIV RNase H and IN and binding mode of 20 in the IN catalytic core (PDB code 3S3M).

3-hydroxyquinazoline-2,4(1H,3H)-diones were also effective inhibitors of HIV-1 RNase H and IN strand transfer activities at sub to low micro­ molar concentrations. Among them, compound 21 (Figure 9.9) was the most promising one with potent inhibitory activity against HIV-1 RNase H (IC50 = 0.41 µM) [40]. This molecule was also a potent IN inhibitor, showing an IC50 value of 0.85 µM. These results reveal the potential of the 3-hydroxyquinazoline-2,4(1H,3H)-dione chemotype as a scaffold to develop potent HIV-1 RNase H and IN inhibitors. A O S N O H 21

H N

B ASP-549

O N OH

ASP-128 GLU-478 HIS-539 ASP-498

O

Dual inhibitor of HIV RNase H and IN

ASP-185 GLU-221

LYS-540

SER-499 TYR-212

Figure 9.9  Chemical structure of compound 21 and its binding modes in the HIV-1 RNase H active site (A, PDB code: 5J1E) and the IN active site (B, PDB code: 3OYA).

282  Advances in Metallodrugs

9.2.5 Inhibitors Obtained by Using Privileged Fragment-Based Libraries The privileged substructure-based diversity-oriented synthesis (pDOS) strategy has proven to be a fruitful tool to rapidly discover biologically active lead compounds by exploring the uncharted chemical space and constructing high-quality compound libraries [41, 42]. For the construction of pDOS libraries, it is crucial to select a privileged substructure with potential for scaffold refining. It is known that hydroxy(iso)quinazoline-2,4(1,3)-dione and its analogs are found in a number of molecules active against a broad spectrum of biological activities. Among them, 2-hydroxy-isoquinoline-1,3(2H,4H)-dione and HPD derivatives are considered the core structures in many antiviral agents. Examples are HIV-1 RNase H active-site inhibitors (compounds 22 and 23) [43] and the hepatitis C virus (HCV) NS5B polymerase inhibitor 24 (Figure 9.10) [44]. The presence of a terminal biaryl substituent seems to be important to increase the efficiency of those compounds. A small library of pyridopyrimidinone derivatives was obtained by modification of the 1-hydroxypyrido[2,3-d]pyrimidin-2(1H)-one scaffold with copper(I)-catalyzed azide-alkyne cycloaddition (CuAAC) reactions [45]. The most active compound of this series, 25 (Figure 9.11) showed

HO

N

HN

O N

O HO

N

N

N O HO O Hydroxypyrimidine, 22 GSK pyridopyrimidinone, 23 HIV RNase H inhibitors

F

OH N N

O N N N

N NH

O

OH N N

O

N

O

O

NS5B, 24 HCV inhibitor

GLU-478 ASP-443

ASP-498

N

GLN-500

O HN

ASP-549 HIS-539

25

26

CN

LYS-540

HIV RNase H inhibitors

Figure 9.10  Representative antiviral agents (22–26) and the binding mode of 26 in the HIV-1 RNase H active site (PDB code: 5J1E).

Molecular HIV-1 and HBV RNase H Inhibitors  283 O

HO

OH O

N H

HO

OH

HO

27

28 OH O

OH

HO

OH

OH

OH O OH

HO

29

OH

30 HIV RNase H inhibitors

Figure 9.11  Chemical structures of HIV RNase H inhibitors 27–30.

remarkable and selective potency against HIV-1 IN. Further studies involving a 5-hydroxypyrido[2,3-b]pyrazin-6(5H)-one scaffold and a variety of substituents led to the identification of compound 26 that exhibited similar inhibitory activity against HIV-1 RNase H and IN strand transfer activities with IC50 values of 1.77 µM and 1.18 µM, respectively [46]. The predicted binding mode of compound 26 in the RNase H active site suggests an interaction between the chelating core and the two divalent catalytic metal cations, which are coordinated to catalytic residues at positions 443, 478, 498, and 549. The hydroxyl group at position 5 and the nitrogen atom at position 4 of 26 can make hydrogen bond interactions that can position 26 close to the active site of the enzyme. Besides, additional hydrogen bonding can be established between the CN group at the terminal of biaryl substituent and the amino group of Lys540 in the HIV-1 RNase H domain, also essential for inhibition. The flexible NH moiety at position 8 of the scaffold introduces conformational flexibility and allows the establishment of π-π interactions between the biphenyl moiety of 26 and the imidazole ring of the highly conserved His539. This interaction restricts the conformational flexibility of His539 and therefore has an influence on the catalytic efficiency of the RNase H.

9.2.6 RNase H Inhibitors in Natural Products Natural extracts have been investigated while searching for innovative chemotypes valuable against many targets. This approach has been applied to HIV-1 RNase H while searching for natural extracts with promising characteristics [47].

284  Advances in Metallodrugs In 2017, Sonar et al. found that extracts of leaves of Ocimum sanctum L. contained molecules with RNase H inhibitory activity. The active compound was a ferulaldehyde containing a catechol moiety that showed activity in the low micromolar range in enzymatic assays (IC50 = 2.4 µM) [48]. Derivatives of that compound included N-oleylcaffeamide (27) that showed remarkable inhibitory activity against HIV-1 RNase H and RNAdependent DNA polymerase activities with IC50 values of 0.68 µM and 2.3 µM, respectively [49]. Prenylated phloroglucinols (28–30, Figure 9.11) obtained from the angiosperm Hypericum scruglii (a plant related to the common Saint John’s wort) were shown to inhibit HIV-1 replication with EC50 values of 3.5 to 8 µM, seven to fourteen times lower than their CC50 [50]. One of these compounds (30) inhibited HIV-1 IN, RNase H and DNA polymerase activities in the micromolar range.

9.2.7 Drug Repurposing Based on Privileged Structures Drug repurposing which consists of giving old inhibitors a new use (outside the scope of the original medical indication) by exploring new molecular pathways and targets for intervention has recently become very popular [51–53]. It has been demonstrated that many compounds selected for the ability to suppress HIV RNase H activity also inhibit HBV RNase H in enzymatic assays. Some of them can also block HBV replication in cell cultures via suppression of the viral RNase H activity [19, 54]. N-hydroxyimides show anti-HBV RNase H activity while exerting potential anti-HIV activity. Representatives of this group of compounds include N-hydroxyisoquinolinediones (HID) (31), HPD (32), and Nhydroxynapthyridinones (HNO) (33). Compounds 31, 32, and 33 have been

N-hydroxyisoquinolinedione (HID)

Cl

N-hydroxynapthyridinone (HNO)

NH2

H N

O N H

N-hydroxypyridinediones (HPD)

N O 31

N O OH

NH2

HO

N O OH

32

N

O N OH 33

HBV RNase H inhibitors

Figure 9.12  Representative N-hydroxyimides (31–33) as HIV/HBV RNase H inhibitors.

Molecular HIV-1 and HBV RNase H Inhibitors  285 characterized as HBV RNase H inhibitors in replicon assays, with EC50 values of 1.4, 0.69, and 3.4 μM, respectively [19, 57] (Figure 9.12). N-hydroxyimide cores provide different directionalities for branching, and preliminary SAR for different substituents and molecular cores have been empirically determined. HID and HPD compounds inhibit the HIV RNase H activity in vitro through binding the two Mg2+ cations in the RNase H active site. The inhibitory effects of HBV replication for HID and HPD derivatives has been measured by using oligonucleotide-directed RNA cleavage assays as well as cell-based HBV replication assays [55]. Both chemotypes (HID and HPD) have been identified as attractive scaffolds for antiviral development [54, 56]. Recently, a preliminary SAR for HPD and HID derivatives was reported [57]. The six-membered nitrogenous HPD ring appears to be the minimal pharmacophore because it is shared by all active compounds (Figure 9.13). Addition of a second six-membered ring bridging positions five and six of the HPD ring creates the HID scaffold. Both the HPD and HID scaffolds have an oxygen trident at positions C1, N2, and C3 which is essential for their activity. The loss of any one of these oxygens results in inactive compounds. For example, compound 34 lacks the oxygen at N2 and is inactive (Figure 9.13). This is consistent with the known mechanism by which the HIDs inhibit the HIV RNase H [58]. The six-membered ring is the minimum ring necessary for activity. Compound 35 that contains a five-membered ring is also inactive, presumably due to an inability to coordinate the Mg2+ ions at the active site of the HBV RNase H. This failure could be due to inappropriate bond angles and/or to the lack of a relative acidic hydrogen at R4 of the HID ring. O

O O

N O NH2

O

O 35

34

N O OH

N H

N O OH

O

5

6 1

2N 3 O

6

H N

7

O

O

8

N O

37

N O OH 38

4

OH HPD

36

F

O O

N

N OH

F

O

5

O 4

1 2N

3

O

OH HID

HBV RNase H inhibitors

Figure 9.13  Compounds (34–38) with anti-HBV potency and their preliminary SAR.

R

286  Advances in Metallodrugs Substitutions by alkyl groups at C4 are not tolerated. However, significant anti-HBV activity (EC50 = 1.4 ± 0.3 μM) can be achieved by including a carbonyl group (ester or amide function) at C4 (38), which increases the acidity of the hydrogen linked to C4 and therefore the ability to coordinate the Mg2+ ions [59], followed by large hydrophobic groups (aryl or alkaryl).

9.3 Conclusion The effectiveness of current antiviral treatments is threatened by the emergence of drug resistance and new drugs will be important to treat HIV/ HBV infections caused by transmitted drug-resistant strains. In this context, drugs acting on novel targets or having stronger inhibitory effects will be helpful to develop more effective treatments. In this chapter, we have covered progress recently made to inhibit the activity of HIV/HBV RNase H using small molecules, while focusing on drug discovery strategies and mechanisms of action of the identified inhibitors. As described above, the vast majority of HIV/HBV RNase H inhibitors act by coordinating to the catalytic center metal ions. The most potent RNase H inhibitors identified for both HIV-1 and HBV are active site chelating molecules containing a ring with fixed angles, which proved to be the best option to achieve inhibition of the enzymatic function. From a pure speculative point of view, given the promising data on HBV RNase H inhibitors, the development of compounds able to block the two viruses at once seems also a really appealing possibility to treat HIV-1/HBV co-infections. It has been reported that the HBV RNase H inhibitors may help improve treatment efficacy enough to clear the virus from the liver when used in combination with existing anti-HBV drugs and/or with other novel inhibitors under development [60]. However, most of the existing HBV RNase H inhibitors were discovered by screening libraries of compounds similar to those used in the identification of many HIV RNase H inhibitors. Expanding this repertoire will require exploring novel chemical spaces. Unfortunately, HTS is limited in part by difficulties in obtaining reliable and relatively high amounts of recombinant HBV RNase H. Although many highly active RNase H active site inhibitors have been described in this section, neither of those molecules is currently moving into advanced preclinical development. Despite their relatively efficiency in vitro, those compounds show only modest effects when tested ex vivo or in animal models. This may be due to poor membrane permeability and cell uptake (probably due in part to the presence of several hydroxyl groups),

Molecular HIV-1 and HBV RNase H Inhibitors  287 poor solubility, or other physicochemical limitations. Nevertheless, the in vitro activity of those compounds indicates that they could become good candidates for development into prodrugs, probably by introducing lipophilic groups to improve their cell permeability. Nevertheless, potent, selective, and orally active HIV and HBV RNase H metal chelating inhibitors to treat those viral infections still have good perspectives for future development.

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Molecular HIV-1 and HBV RNase H Inhibitors  289 efavirenz and/or a β-thujaplicinol analogue. Biochem. J., 455, 179–184, 2013. 27. Lu, G., Lomonosova, E., Cheng, X., Moran, E.A., Meyers, M.J., Le Grice, S.F., Tavis, J.E., Hydroxylated tropolones inhibit hepatitis B virus replication by blocking viral ribonuclease H activity. Antimicrob. Agents Chemother., 59, 1070–1079, 2015. 28. Hu, Y., Cheng, X., Cao, F., Huang, A., Tavis, J.E., β-Thujaplicinol inhibits hepatitis B virus replication by blocking the viral ribonuclease H activity. Antiviral Res., 99, 221–229, 2013. 29. Tang, J., Liu, F., Nagy, E., Miller, L., Kirby, K.A., Wilson, D.J., Wu, B., Sarafianos, S.G., Parniak, M.A., Wang, Z., 3-Hydroxypyrimidine-2,4-diones as selective active site inhibitors of HIV reverse transcriptase-associated RNase H: Design, synthesis, and biochemical evaluations. J. Med. Chem., 59, 2648–2659, 2016. 30. Ju, H., Zhang, J., Huang, B., Kang, D., Huang, B., Liu, X., Zhan, P., Inhibitors of influenza virus polymerase acidic (PA) endonuclease: Contemporary developments and perspectives. J. Med. Chem., 60, 3533–3551, 2017. 31. Kankanala, J., Kirby, K.A., Liu, F., Miller, L., Nagy, E., Wilson, D.J., Parniak, M.A., Sarafianos, S.G., Wang, Z., Design, synthesis, and biological evaluations of hydroxypyridonecarboxylic acids as inhibitors of HIV reverse transcriptase associated RNase H. J. Med. Chem., 59, 5051–5062, 2016. 32. Kolb, H.C., Finn, M.G., Sharpless, K.B., Click chemistry: Diverse chemical function from a few good reactions. Angew. Chem. Int. Ed., 40, 2004–2021, 2001. 33. Takayama, Y., Kusamori, K., Nishikawa, M., Click chemistry as a tool for cell engineering and drug delivery. Molecules, 24, e172, 2019. 34. Pala, N., Esposito, F., Rogolino, D., Carcelli, M., Sanna, V., Palomba, M., Naesens, L., Corona, A., Grandi, N., Tramontano, E., Sechi, M., Inhibitory effect of 2,3,5,6-Tetrafluoro-4-[4-(aryl)-1H-1,2,3-triazol-1-yl]benzenesulfonamide derivatives on HIV reverse transcriptase associated RNase H activities. Int. J. Mol. Sci., 17, e1371, 2016. 35. Nowotny, M., Retroviral integrase superfamily: The structural perspective. EMBO Rep., 10, 144–151, 2009. 36. Corona, A., Leva, F.S.D., Thierry, S., Pescatori, L., Crucitti, G.C., Subra, F., Costi, R., Identification of highly conserved residues involved in inhibition of HIV-1 RNase H function by diketo acid derivatives. Antimicrob. Agents Chemother., 58, 6101–6110, 2014. 37. Cuzzucoli Crucitti, G., Métifiot, M., Pescatori, L., Messore, A., Madia, V.N., Pupo, G., Saccoliti, F., Scipione, L., Tortorella, S., Esposito, F., Corona, A., Cadeddu, M., Marchand, C., Pommier, Y., Tramontano, E., Costi, R., Di Santo, R., Structure-activity relationship of pyrrolyl diketo acid derivatives as dual inhibitors of HIV-1 integrase and reverse transcriptase ribonuclease H domain. J. Med. Chem., 58, 1915–1928, 2015. 38. Costi, R., Métifiot, M., Chung, S., Cuzzucoli Crucitti, G., Maddali, K., Pescatori, L., Messore, A., Madia, V.N., Pupo, G., Scipione, L., Tortorella, S.,

290  Advances in Metallodrugs Di Leva, F.S., Cosconati, S., Marinelli, L., Novellino, E., Le Grice, S.F., Corona, A., Pommier, Y., Marchand, C., Di Santo, R., Basic quinolinonyl diketo acid derivatives as inhibitors of HIV integrase and their activity against RNase H function of reverse transcriptase. J. Med. Chem., 57, 3223–3234, 2014. 39. Wu, B., Tang, J., Wilson, D.J., Huber, A.D., Casey, M.C., Ji, J., Kankanala, J., Xie, J., Sarafianos, S.G., Wang, Z., 3-Hydroxypyrimidine-2,4-dione-5-Nbenzylcarboxamides potently inhibit HIV-1 integrase and RNase H. J. Med. Chem., 59, 6136–6148, 2016. 40. Gao, P., Cheng, X., Sun, L., Song, S., Álvarez, M., Luczkowiak, J., Pannecouque, C., De Clercq, E., Menéndez-Arias, L., Zhan, P., Liu, X., Design, synthesis and biological evaluation of 3-hydroxyquinazoline-2,4(1H,3H)-diones as dual inhibitors of HIV-1 reverse transcriptase-associated RNase H and integrase. Bioorg. Med. Chem., 27, 3836–3845, 2019. 41. Oh, S. and Park, S.B., A design strategy for drug-like polyheterocycles with privileged substructures for discovery of specific small-molecule modulators. Chem. Commun. (Camb.), 47, 12754–12761, 2011. 42. Kim, J., Kim, H., Park, S.B., Privileged structures: Efficient chemical “navigators” toward unexplored biologically relevant chemical spaces. J. Am. Chem. Soc., 136, 14629–14638, 2014. 43. Vernekar, S.K., Liu, Z., Nagy, E., Miller, L., Kirby, K.A., Wilson, D.J., Kankanala, J., Sarafianos, S.G., Parniak, M.A., Wang, Z., Design, synthesis, biochemical, and antiviral evaluations of C6 benzyl and C6 biarylmethyl substituted 2-hydroxylisoquinoline-1,3-diones: Dual inhibition against HIV reverse transcriptase-associated RNase H and polymerase with antiviral activities. J. Med. Chem., 58, 651–664, 2015. 44. Chen, Y.L., Tang, J., Kesler, M.J., Sham, Y.Y., Vince, R., Geraghty, R.J., Wang, Z., The design, synthesis and biological evaluations of C-6 or C-7 substituted 2-hydroxyisoquinoline-1,3-diones as inhibitors of hepatitis C virus. Bioorg. Med. Chem., 20, 467–479, 2012. 45. Gao, P., Zhang, L., Sun, L., Huang, T., Tan, J., Zhang, J., Zhou, Z., Zhao, T., Menéndez-Arias, L., Pannecouque, C., De Clercq, E., Zhan, P., Liu, X., 1-Hydroxypyrido[2,3-d]pyrimidin-2(1H)-ones as novel selective HIV integrase inhibitors obtained via privileged substructure-based compound libraries. Bioorg. Med. Chem., 25, 5779–5789, 2017. 46. Sun, L., Gao, P., Dong, G., Zhang, X., Cheng, X., Ding, X., Wang, X., Daelemans, D., De Clercq, E., Pannecouque, C., Menéndez-Arias, L., Zhan, P., Liu, X., 5-Hydroxypyrido[2,3-b]pyrazin-6(5H)-one derivatives as novel dual inhibitors of HIV-1 reverse transcriptase-associated ribonuclease H and integrase. Eur. J. Med. Chem., 155, 714–724, 2018. 47. Tatyana, K., Francesca, E., Luca, Z., Giovanni, F., Cheng, Y.C., Ginger, E.D., Enzo, T., Inhibition of HIV-1 ribonuclease H activity by novel frangula-emodine derivatives. Med. Chem., 5, 398–410, 2009. 48. Sonar, V.P., Corona, A., Distinto, S., Maccioni, E., Meleddu, R., Fois, B., Floris, C., Malpure, N.V., Alcaro, S., Tramontano, E., Cottiglia, F., Natural

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10 The Role of Metals and Metallodrugs in the Modulation of Angiogenesis Mehmet Varol1* and Tuğba Ören Varol2 Department of Molecular Biology and Genetics, Faculty of Science, Kotekli Campus, Mugla Sitki Kocman University, Mugla, Turkey 2 Department of Chemistry, Faculty of Science, Kotekli Campus, Mugla Sitki Kocman University, Mugla, Turkey 1

Abstract

Drug development and discovery studies have a great importance because of the presence of many deadly diseases without convenient medical cures available. Thus, substantial research efforts and financial supports have been consumed to have successful drugs and treatment strategies. Metals and metal-based materials have an essential significance in medicinal chemistry. Cancer is the most attractive research subject for pharmacologists, but there are more than 200 different types of cancer and a cancer tissue composes of diverse cancer cells that have different mutations, epigenetic profiles and characters. It seems to be therefore more rational and beneficial to target the hallmarks of carcinogenesis for development and discovery of new treatment strategies and drugs. As one of the most important cancer hallmarks, angiogenesis is defined as the formation of new capillaries from the existing blood vessels. The cells forming tumor tissues require angiogenesis as well as the healthy cells in order to be able to continue their vital activities, to increase the mass and volume of tumor tissue, to sustain numerical increases, to migrate to different regions of the organism. We, therefore, would like to draw the attention of scientists more on the role of metals and metallodrugs in the modulation of angiogenesis. Keywords:  Angiogenesis, cancer, vasculogenesis, metal ions, metallodrugs, metal-based medicine

*Corresponding author: [email protected]; [email protected] Shahid-ul-Islam, Athar Adil Hashmi and Salman Ahmad Khan (eds.) Advances in Metallodrugs: Preparation and Applications in Medicinal Chemistry, (293–318) © 2020 Scrivener Publishing LLC

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10.1 Introduction Sustaining many biological processes and maintaining the human homeostasis within the safe borders that have a subtle balance need the presence of metal ions that play a crucial role as the essential components in the normal functioning of proteins, and operating the completion, stabilization, and regulation courses of the cellular functions [1, 2]. Realizing the importance of metal ions and metal-binding components into the biological systems has drawn the attention of many scientists in the field of medicinal inorganic chemistry, and design and discovery of metal-based complexes have become one of the most attractive research fields for the diagnosis and therapy of many diseases [3, 4]. Moreover, it should be noted that there is an evolutionary transportation and distribution system for metal ions and metal-binding components because of the prominent tasks of metals for the progression of many important cellular events. The presence of sophisticated and sensitive systems for the transportation and distribution of metal ions and metal-binding components have attracted the intense attention of researchers, and the metal complexes have been designed and synthesized by considering their cellular transportation and distribution systems [3, 5, 6]. Although medicinal inorganic chemistry and the medical application of metal-based drugs and remedies seem to be regarded in the modern era, there is a large collection of the ancient sources dated back to the civilizations of India, China, Egypt, and Mesopotamia that provides the earliest reports of the historical use of metals for medicinal purposes [7]. For example, the ancient Egyptian medical treatise that is known as the Edwin Smith Papyrus has some explanations for the medicinal use of copper to sterilize chest wounds [8]. It is also well known that gold, silver, and arsenic were employed in ancient medicine [9­–12]. The gold-based remedies were used to treat diseases as far back as 2500 BC in China, and silver was applied to treat ulcers and wound by Greek physician Hippocrates [9–12]. Arsenic was employed as escharotic in the form of realgar (As4S4) and orpiment (As2S3) to remove the unwanted or diseased tissues by a number of predominant physicians including Hippocrates [9–12]. Apart from the historical use of arsenic, it has shown up again in modern medicine as the first therapeutic metallodrug named “salvarsan” (Figure 10.1) that is a mixture of 3-amino-4-hydroxyphenyl-arsenic (III) [3]. In 1912, the German Nobel laureate Paul Ehrlich discovered salvarsan (arsphenamine) to treat the sexually transmitted infection named syphilis, which is caused by the bacterium Treponema pallidum [13–15]. Ehrlich

Metals and Metallodrugs in Angiogenesis  295 N H2

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Figure 10.1  Salvarsan (Arsphenamine) was firstly known to have As=As double bond, but the mass spectral studies showed in 2005 that the drug salvarsan is a mixture consisting of cyclo-As3 and cyclo-As5 species. Neosalvarsan was synthesized by the modification of salvarsan in the laboratory of Paul Ehrlich to obtain a water-soluble, lesstoxic, and active analog of salvarsan.

pioneered scientific concept named “Magic Bullet”, and hypothesized that it is possible to fight and kill the invading microorganisms without any damage in the host organisms by using a convenient drug that is specific to the target microorganisms [13–15]. Although the exact composition of salvarsan is still obscure, salvarsan and neosalvarsan (less toxic, but more water-soluble version of salvarsan) (Figure 10.1) had been widely

296  Advances in Metallodrugs employed as a standard remedy to treat syphilis by adding mercury and bismuth as far as the discovery of penicillin after World War II [3, 16]. However, it should be noted that the Magic Bullet concept along with the development of arsenic-based drugs provided a rational approach to drug design and precisely placed in the field of medicinal inorganic chemistry as a significant milestone [7, 17]. In addition to the inspiring study of Paul Ehrlich, it should be remembered and underlined the contribution of the Swiss chemist Alfred Werner, who has been known as the founder of coordination chemistry [18, 19]. Werner was awarded the Nobel Prize for Chemistry in 1913 thanks to “the recognition of his work on the linkage of atoms in molecules by which he has thrown new light on earlier investigations and opened up new fields of research especially in inorganic chemistry” [18, 19]. The establishment and development of coordination chemistry as a different and novel discipline of chemistry opened up new research fields such as the medicinal metal coordination chemistry that focuses on the design and synthesis of metal-based compounds in therapy and diagnosis [19, 20]. Biological activity of metal-based compounds had been understood along with the presence of the Werner’s coordination theory, which clarifies that the activity of metals is not only depended on the type of metals and the oxidation state but also the coordination geometry and the number and type of coordinated ligands [19, 21, 22]. Apart from the significant contribution of Alfred Werner in the medicinal metal coordination chemistry, he gave a special importance to the platinum complexes to understand their chemical structures and behaviors by studying between 1893 and 1920, and publishing 14 papers [23]. Werner’s studies about platinum complexes lead to put forth the chemical structure and behavior of cis-diamminedichloroplatinum(II) (cisplatin) by displaying that ammonia can donate its lone pair of electrons to a metal ion such as platinum (II) in a dative or coordinate bond [24, 25]. Cisplatin was known as “Peyrone’s salt” because it was firstly discovered and synthesized in 1845 by the Italian chemist Michele Peyrone [24, 25]. The structure of cisplatin could not be elucidated until Alfred Werner discovered its sterical configurations [7]. Cisplatin can be synthesized by the reaction of potassium tetrachloroplatinate with ammonia, and a square planar molecule with three chlorides and one ammonia, which was displaced by one of the four chlorides. After the addition of first ammonia, the second ammonia replaces the chloride ligand adjacent to the first ammonia because chloride has a stronger trans effect than ammonia [24–26]. After approximately 70 years from the elucidation of cisplatin structure and behavior, Barnett Rosenberg and coworkers serendipitously discovered the biological activity of the square planar platinum (II) complex cisplatin

Metals and Metallodrugs in Angiogenesis  297 and its platinum (IV) analog on the bacteria named Escherichia coli [1, 27, 28]. The study was published in Nature by the title “Inhibition of cell division in Escherichia coli by electrolysis products from a platinum electrode” [1, 27, 28]. After a series activity studies, Rosenberg and coworkers hypothesized that cisplatin and a number of analogs might inhibit the fast growing cancer cells [1, 26, 28]. The studies performed by Rosenberg and coworkers showed that these metal complexes inhibit the development of the solid sarcoma 180 [1, 26, 28]. Thus, the era of metallodrugs was opened up for their anticancer activity applications [1, 26, 28].

10.2 Metallodrugs in Anticancer Therapy After the anticancer activity of cisplatin (Platinol) was investigated by Rosenberg and coworkers, it was patented by U.S. Patent Office, and approved in December 1978 by the Food and Drug Administration [28–33]. As the first approved and the most successful therapeutic metallodrug, cisplatin (Figure 10.2) was employed alone or in combination with other drugs to treat various types of human malignancies such as head and neck, lung, ovarian, testicular and bladder cancers [1, 3, 34]. Cisplatin is still considered as an important anticancer drug that has a prominent place in the modern oncology because it is employed as a standard anticancer therapy against many types of cancer, such as anal, bladder, brain, cervical, ovarian, testicular, and non-small cell lung cancer [3, 35]. The achieved success of cisplatin by the wide range application on many types of cancer and the high annual sales rate with ca. US$500 million drawn the attention of scientists on metallodrugs and their anticancer potentials [1, 34]. Thus, the studies about metal-based drugs in the pharmaceutical industry have been dramatically increased by designing, synthesizing and testing several thousand metallodrug analogs to find more potent anticancer drugs [36, 37]. As a result of the comprehensive research, second-, third-, and even fourth-generation platinum analogs have been submitted to receive the approval of the Food and Drug Administration and other national authorities [1]. Carboplatin (Paraplatin) and oxaliplatin (Eloxatin) (Figure 10.2) have been approved by the Food and Drug Administration, though nedaplatin (Aqupla), lobaplatin (D-19466), heptaplatin (SKI-2053R), and miriplatin (Miripla) (Figure 10.2) have received limited approval from the national authorities, respectively, in Japan, China, South Korea, and Japan [34]. Although carboplatin has been clinically used in worldwide for a broad range of solid neoplasms, oxaliplatin is employed for metastatic colorectal cancer [34, 38, 39]. Nedaplatin is used for several solid neoplasms

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Picoplatin

Figure 10.2  Platinum-based anticancer metallodrugs approved worldwide, approved by the national authorities and in clinical trials.

Metals and Metallodrugs in Angiogenesis  299 in Japan; lobaplatin is employed to treat inoperable, small cell lung and metastatic breast cancer, and chronic myelogenous leukaemia (CML) in China; heptaplatin is applied for advanced gastric cancer in South Korea; and miriplatin is used for hepatocellular carcinoma in Japan [34, 38, 39]. On the other hand, there are some other platin-based drugs such as lipoplatin (liposomal cisplatin), ProLindac (AP5346; diaminocyclohexane platinum polymer prodrug), picoplatin, and satraplatin (Figure 10.2) that are in the various phases of clinical development to treat different types of cancer [34, 36, 40–44]. It is clear from the literature that there is a considerable publication mass about the metal-based anticancer drugs, though the underlying molecular mechanisms of their toxicological and pharmacological profiles seem to be not fully elucidated [34]. It has been, for example, supposed for a long while that cisplatin display activity by aquation inside cells because of the varying Cl− concentration [45, 46]. However, platinum(II)-based drugs include [Pt(NH3)2]2+ unit, and Pt2+ of this unit binds covalently to the N-7 of adenine or guanine bases in the dinucleotide sequences AG and GG to form cisplatin−DNA adducts [47–49]. Cisplatin−DNA adducts cause changes in DNA structure, leading replication arrest and transcription inhibition, which ultimately result in the induction of DNA repair mechanisms, cell-cycle arrests, and apoptosis [47–49]. Apart from the needs of pharmacological and toxicological mechanism studies, it has been observed that the platinum-based anticancer agents have some serious limitations such as having side effects like myelosuppressive effects, nephropathy, nephrotoxicity, ototoxicity, neutropenia, diarrhea, vomiting, etc., and restraining because of the drug resistance phenomenon after recurrent chemotherapy [1, 39, 50–52]. Due to the prominent limitations of the metallodrugs on the market today, many medicinal chemists have tried to develop encapsulated or polymer- and biomolecule-integrated forms of the approved metallodrugs such as lipoplatin (liposomal cisplatin) and non-platinum-based drugs such as gold, ruthenium, indium, titanium and gallium-based drugs [5, 6, 53–57]. Moreover, the drug delivery strategies have gained more importance to improve water solubility, circulation time and selective transportation in the human body, specific tumor delivery, controlled release, and long-term anticancer efficacy of metallodrugs [37, 58–61]. Apart from the drug design, development, and synthesis studies, new treatment modalities and combinational therapy strategies that use the conventional drugs have been proposed to improve the clinical outcomes [62]. As widely known, the most of the chemotherapeutics such as the approved platinum-based drugs, cisplatin, carboplatin, and oxaliplatin, have been

300  Advances in Metallodrugs conventionally applied at the maximum tolerated dose to destruct the maximum possible number of cancer cells [63, 64]. However, it was found that the conventional application of the chemotherapy by using high and intense dose in a short treatment period have a key role in the formation of drug resistance, systemic toxicity, and potential long-term side effects [62, 65]. The novel treatment strategies such as metronomic therapy and adaptive therapy were therefore emerged with better clinical outcomes, though the conventional drugs have been employed [66–68]. The purpose of the metronomic cancer therapy is defined as killing the maximum number of cancer cells with a greater cumulative amount of the anticancer agent that can be loaded into the body by employing low dose of anticancer agent with no prolonged drug-free breaks [62]. The purpose of the adaptive therapy (the eco-evaluation-based chemotherapy) is defined as limiting the tumor size and proliferation of the drug resistant and highly aggressive cancer cell population [62]. These aggressive cancer cells are located into the center of tumor tissue and can be kept under control thanks to the surrounding drug sensitive cancer cells by employing low dose of anticancer agent with frequent application [62]. Despite the development of metal-based anticancer drugs and the novel metallodrug-based treatment strategies, it is explicitly observed that the cytotoxic treatment has approached always provoke more or less side effects [1, 62]. Additionally, it should be noted that we do not have yet any drug that has comprehensive cytotoxic activity on the wide variety of cancer diseases, even if it is ignored that a cancerous tissue is formed by morphologically, functionally, and biologically heterogeneous texture comprised various cancer cells with different characteristics, epigenetic profiles, and mutations [69, 70]. The attraction of the scientist has been therefore drawn to the common features of carcinogenesis to improve more rational and beneficial treatment strategies, and angiogenesis is considered as most important target to treat cancers [70–74].

10.3 Angiogenesis as a Substantial Target of Tumorigenesis As one of the most important cancer hallmarks, angiogenesis that can be simply described as the formation of new capillaries from the existing blood vessels plays a crucial role in carcinogenesis by providing oxygen to the cancer cells and removing the metabolic wastes [70]. Additionally, the cancer cells have an opportunity to migrate into the vascular system and metastasize thanks to the tumor angiogenesis [70, 75, 76]. Angiogenesis

Metals and Metallodrugs in Angiogenesis  301 should be therefore considered as a substantial target to inhibit cancer tissue growth and metastasis [70, 75, 76]. Although angiogenesis is a normal physiological and biological process for reproduction, healing, tissue growth, the constitution of endometrium during the menstrual cycle and the development of fetus during pregnancy, tumor angiogenesis is an abnormal physiological and biological process and lead to the formation of abnormal capillary vessels [77–80]. The abnormal capillaries are characterized with the leakages, ruptures, vessel dilation, irregular vascular networks, abnormal morphology of endothelial cells, and vascular mimicry [77–80]. Vascular mimicry can be described as the ability of cancer cells to have endothelial cell-like properties, behave like endothelial cells and form the vascular pattern along with the host endothelial cells [77–80]. A cell should be located at a maximum distance of 100–200 µm from the capillary vessel to achieve sufficient oxygen, since the diffusion limit of oxygen within the tissue is 100–200 µm [81]. In normal physiological conditions, there is a balance between the tissue growth and angiogenesis, and the balance is regulated by angiogenesis inducing (angiogenic) factors such as vascular endothelial growth factor (VEGF), epidermal growth factor (EGF), fibroblast growth factor (FGF), angiopoietins, integrins, transforming growth factor beta (TGF-beta) and matrix metalloproteinase (MMP) enzymes, and angiogenesis suppression (anti-angiogenic) factors such as tissue factor pathway inhibitor (TFPI), angiostatin, endostatin, osteopontin, collagen, kininogens, trombospondin-1 and -2, angiopoietin-2, and platelet factor-4 [70, 82–86]. The balance between the angiogenic and anti-angiogenic factors provides an angiogenic switch, and the opening and closing of this switch regulates the formation of new blood vessels [70, 87]. When the cancer cells, for example, are stayed away from the capillary vessels because of the uncontrolled tumor tissue growth, the oxygen deprived cancer cells (hypoxic cancer cells) secrete angiogenic factors, open the angiogenic switch and induce angiogenesis, and so new capillary vessels are formed within the tumor tissue [70, 82]. New capillary vessels can be formed in utero and adults by two different types of angiogenesis that are named as “sprouting angiogenesis” and “intussusceptive angiogenesis” [88–91]. Sprouting angiogenesis is known as a more complicated process that includes the debilitation of cell-cell interactions, capillary basement membrane degradation, endothelial cell proliferation, extensive cell migration, tubulogenesis (the constitution of capillary like structures), vessel fusion (anastomosis of the new tubular sprouts), constitution of the new basement membrane, and the functional contribution of pericytes [90–94]. The intussusceptive angiogenesis, called also as splitting angiogenesis, is categorized as rapid, efficient, and

302  Advances in Metallodrugs more economic process in the metabolic and energetic manners because it only needs reorganization of the endothelial cells that are already located into the existing capillaries to form new capillary vessels, which are therefore less leaky than the sprouting capillary vessels [90–94]. Therefore, the anti-angiogenic drug candidates should target the common features of these two angiogenesis processes such as tubulogenesis, vessel fusion, and angiogenic factors rather than the endothelial cell proliferation, invasion, and migration.

10.4 Metals and Metallodrugs in Angiogenesis Some unwanted effects of metal-based drugs such as drug resistance, systemic toxicity and potential long-term side effects have compelled the scientist to design and synthesize novel metal-based anticancer drug candidates by using different metals such as ruthenium, silver, copper, gold, etc. [62]. It seems that gold, copper, silver, platinum, ruthenium, zinc, and cadmium complexes, respectively, are the most studied metal-based nanoparticles and complexes as angiogenesis modulators though the literature about the angiogenesis modulation activity of metallodrugs seems to be very poor and urgently needs to be expended. Apart from the common metals, there are some interesting studies about chromium, neodymium, iridium, magnesium, rhodium, arsenic, and cerium complexes [95–101]. One of these interesting studies was published by Duraipandy and coworkers in 2019, the study has indicated that the rare earth metal, neodymium, nanoparticles induce intracellular ROS generation via PKM2-NOX4 signaling pathways depending on the shape of the neodymium nanoparticles, and the ROS generation was found to be directly correlated with the activation of the pro-angiogenic factors such as HIF1-α (hypoxia inducible factor 1-α), VE-cadherin (vascular endothelial cadherin), VEGF, and VEGFR-2 to promote angiogenesis [97]. In another study, the angiogenesis modulation activities of biodegradable magnesium (Mg) alloys have been investigated because of the usage of magnesium-based stents in cardiovascular applications, and it was found that Mg ions affect endothelial cell proliferation, migration, cytoskeletal reorganization, and the gene expression related to the angiogenesis and cell adhesion signaling pathways [99]. Moreover, it was reported that cerium oxide nanoparticles and cyclometalated rhodium(III) complex inhibit angiogenesis [100, 101]. On the other hand, copper-based nanoparticles and complexes have been emerged as one of the most studied metal-based angiogenesis modulators because copper ions have an ability to stimulate angiogenesis by regulating

Metals and Metallodrugs in Angiogenesis  303 the expression levels of VEGF, FGF-2, and angiogenin in endothelial cells [102–106]. However, it is also well known that the excess copper in the cells can show a toxic effect by inducing ROS generation and JNK phosphorylation, leading to caspase activation [102–106]. Thus, the activities of copper ions and copper-bearing alloys on endothelial cells and angiogenesis have been investigated, especially for the cardiovascular queries, and copper-bearing metal stents increase endothelial cell proliferation and angiogenesis [107, 108]. Some of the copper-based materials such as copper complex of quercetin, hydrocolloid of copper nanoparticles, and copper-sulphated hyaluronic acid complex have been identified as angiogenesis stimulators [109–111]. The other anti-angiogenic copper-based materials have been reported as copper oxide nanoparticles, the histidine– proline rich glycoprotein-copper complex, cetuximab-modified copper(II) sulfide nanoparticles, mononuclear copper(II) polypyridyl complexes, and copper-schiff base complexes [102, 112–116]. Gold-based nanoparticles and complexes emerge as the most studied metal-based angiogenesis modulators, but they have been generally considered as angiogenesis inhibitors except in a very small number of studies [117]. It was, for example, proposed in the literature that nanoconjugates containing chloroauric acid (HAuCl4) and the pro-angiogenic leaf extract of Hamelia patens, and nanocomposites containing type I collagen and gold nanoparticles act as angiogenesis stimulators and may be employed for the vascular regeneration [118, 119]. Moreover, Roma-Rodrigues and coworkers (2016) reported that gold nanoparticles can be used as ideal theranostic vehicles and the formulation of gold nanoparticles with angiogenic peptides may be employed for revascularization [120]. However, the vast majority of the studies about gold-based angiogenesis modulators is about their negative modulation of angiogenesis, and gold-based nanoparticles and complexes generally are considered as anticancer agents along with their anti-angiogenic activities [117]. The studies that were performed by using the bare gold nanoparticles showed that the gold nanoparticles inhibit endothelial cell migration, capillary-like tube formation and angiogenesis by blocking interleukin-6, the function of pro-angiogenic heparinbinding growth factors such as VEGF165 (vascular endothelial growth factor 165) and bFGF (basic fibroblast growth factor), and the interaction between VEGF165 and VEGFR2 [121–124]. The gold nanoparticles have been also used as vehicles to load the therapeutic cargos such as quinacrine, naturally derived phytochemicals (curcumin, turmeric, quercetin, and paclitaxel), anti-angiogenic recombinant human endostatin and captoprilpolyethyleneimine to combine and improve their anti-angiogenic activities [125–128]. Moreover, gold nanoparticles are conjugated and encapsulated

304  Advances in Metallodrugs with some natural products such as aqueous extracts of Mangifera indica seeds, Gum Arabic (acacia gum), snake venom protein toxin NKCT1, and heparin derivative [129–132]. Furthermore, some studies reported novel anti-angiogenic gold-based complexes that were designed, synthesized by using mannose-6-phosphate analogs and boronated phenylalanine derivatives, which can use L-amino-acid-transport system to pass across the cell membrane, and the boronated phenylalanine derivative-based gold complexes were found significantly anti-angiogenic [5, 6, 133]. Besides the boronated phenylalanine derivative-based gold complexes, Varol and coworkers also designed and synthesized the boronated phenylalanine derivative-based platinum complexes using 4,4’-dimethyl-2,2’dipyridyl, 1,10-phenanthroline-5,6-dion, 2, 2’-dipyridyl, and 4, 4’diaminobibenzyl ligands, and they reported that these platinum complexes also have anti-angiogenic activities depending on the type of ligands [5, 6]. Although there are so many studies about the platinum-based anticancer drugs because of the clinical success of cisplatin, there are limited studies about the anti-angiogenic activities of platinum-based drugs. Zamora and coworkers (2017) reported some antivascular organoplatinum(II) complexes, and Yang and coworkers (2016) published a paper about Pt(II) and Pt(IV) phosphaplatins that inhibit VEGFR-2 in HUVEC cells [134–137]. Moreover, Karmakar and coworkers (2016) reported that cis-dichlori­ doplatinum(II) complex of a chelating nitrogen mustard inhibits angiogenesis in chick embryo, and Peterson and coworkers (2017) published a paper about platinum-based chemotherapeutics that have anti-angiogenic activity through glycan targeting [134–137]. It is well known that the growing interest of scientists has been drawn to the non-platinum metallodrugs such as ruthenium, silver, zinc, cadmium, titanium, and cobalt complexes to avoid the unwanted effects of platinum-based drugs such as serious side effects, systemic toxicity, and drug resistance phenomenon and to improve the pharmacological properties [1]. These novel non-platinum complexes were also investigated for their anti-angiogenic activities. For example, ruthenium complexes are considered more beneficial compared to the platinum complexes because of the reduced toxicity, different action mechanisms, non-cross resistance, and different activity spectrum [138]. Additionally, the different research groups have synthesized various ruthenium-based materials such as ruthenium-thiol protected selenium nanoparticles, trans-imidazole dimethyl sulphoxide tetrachloro-ruthenate, potassium chlorido (ethylene diammino tetra acetate) rutenate, sodium (bis-indazole) tetrachlororuthenate, heterometallic ruthenium–gold, ruthenium(II)-arene, ruthenium(II)-p-cymene, and ruthenium(II)-benzimidazole complexes, and

Metals and Metallodrugs in Angiogenesis  305 they generally reported that these ruthenium-based materials have significant anti-angiogenic activities [139–146]. On the other hand, silver-based nanoparticles that have been prepared by green biosynthetic method using different natural products, such as the flowers extract of Achillea biebersteinii and the biomass of Bacillus licheniformis, were identified as antiangiogenic nanoparticles [147–149]. Moreover, the underlying mechanism of anti-angiogenic activity of silver nanoparticles prepared with Bacillus licheniformis has been reported as the targeting of VEGF induced cell survival via PI3K/Akt dependent pathway [147–149]. The poly(lactic-coglycolic acid)-based uniform hybrid nanoparticle (QAgNP) that is prepared with quinacrine and silver, and heparin-conjugated silver nanoparticles that is prepared by reducing silver nitrate with diaminopyridinyl-derivatized heparin (DAPHP) polysaccharides have showed significantly anti-angiogenic activity, especially DAPHP-reduced silver nanoparticles inhibited basic fibroblast growth factor (bFGF-2)-induced angiogenesis [132, 150]. Additionally, Kang and coworkers (2011) reported that polyvinylpyrrolidone (PVP)-coated silver nanoparticles stimulate angiogenesis by regulating endothelial cell tube formation, ROS production, angiogenic factors such as nitric oxide (NO) and vascular endothelial growth factor (VEGF), and VEGF receptor (VEGFR)-mediated signaling factors such as ERK1/2, Akt, FAK and p38 [151]. Zinc-based complexes have been also considered as anti-angiogenic formulations such as rutin-zinc(II) flavonoid-metal complex that down-regulate the expression levels of VEGF and cell cycle-related gene Cyclin D1 [152]. On the other hand, Nethi and coworkers reported in 2017 that zinc-doped titanium oxide nanoparticles have pro-angiogenic properties by inducing the production of proangiogenic messengers such as nitric oxide and reactive oxygen species, and the phosphorylation of STAT3 at Tyr705 residue and p38 MAPK at Thr180/Tyr182 residue [153]. However, pro-angiogenic effects of zincdoped titanium oxide nanoparticles seem to be mainly based on the titanium oxides rather than zinc-based parts [154]. Freyre-Fonseca and coworkers (2018) reported that titanium dioxide nanoparticles dispersed in saline solution and fetal bovine serum have anti-angiogenic effects by down regulation in the pattern of VEGF expression, but the titanium dioxide nanoparticles do not lead any differences on the expression level of VEGF [154]. Similar to the titanium-based nanoparticles, low dose intake of the heavy metal cadmium increases the level of hypoxia inducible factor 1-α (HIF1-α), and that positively regulate the expression levels of VEGF-A and VEGFR2 through ROS, ERK, and AKT signaling pathways though cadmium is considered as a highly toxic and carcinogenic metal [155, 156].

306  Advances in Metallodrugs Some limited studies, especially with the nickel, manganese, iron, bismuth, and selenium focused on the modulation of angiogenesis have been also published by different research groups [116, 157–163]. For example, Couto and coworkers (2016) determined that bismuth subgallate delays the angiogenesis and the mucosal wound healing, and Parrilha and coworkers (2012) reported that lapachol-based bismuth complex has a significant anti-angiogenic activity that is more than the anti-angiogenic potential of free lapachol [164, 165].

10.5 Concluding Remarks and Future Prospects It is clear that metals and metal-based materials such as complexes, nanoparticles, polymers, conjugations, and encapsulations have a great importance and potential for anticancer therapies. Targeting angiogenesis seems to be also one of the most rational treatment strategies because of the cellular heterogeneity of cancer tissues and the wide variety of cancer diseases. However, reviewing the angiogenesis modulation role of metals and metal-based materials demonstrated that the literature should be still excessively expanded because there is a great limitation, especially for the anticancer metal-based drugs, such as the unwanted toxicity on healthy cells, tissues and organs, and this limitation can be eliminated by designing and discovering of the non-toxic but anti-angiogenic metal-based materials. On the other hand, we think that it should be considered to design and synthesize new metal-based materials by combination of already known anticancer and anti-angiogenic drugs to have new successful materials with the improved pharmacological activities and profiles. Developing multidisciplinary research projects about metal-based materials that possibly have angiogenesis modulation activity has a special importance. We consequently think that only different perspectives that are derived from the knowledge of various scientific disciplines such as chemistry, bioinformatics, molecular biology, genetics, medicine, and pharmacology can accelerate to have prosperous results and clinically successful materials.

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316  Advances in Metallodrugs properties and inhibits metastasis and angiogenesis-associated proteases in renal cancer. JBIC J. Biol. Inorg. Chem., 23, 399–411, 2018. 140. Berndsen, R.H., Weiss, A., Abdul, U.K., Wong, T.J., Meraldi, P., Griffioen, A.W., Dyson, P.J., Nowak-Sliwinska, P., Combination of ruthenium (II)arene complex [Ru (η 6-p-cymene) Cl 2 (pta)](RAPTA-C) and the epidermal growth factor receptor inhibitor erlotinib results in efficient angiostatic and antitumor activity. Sci. Rep., 7, 43005, 2017. 141. Bhattacharyya, S., Purkait, K., Mukherjee, A., Ruthenium (ii) p-cymene complexes of a benzimidazole-based ligand capable of VEGFR2 inhibition: Hydrolysis, reactivity and cytotoxicity studies. Dalton Trans., 46, 8539–8554, 2017. 142. Nowak-Sliwinska, P., Clavel, C.M., Păunescu, E., Te Winkel, M.T., Griffioen, A.W., Dyson, P.J., Antiangiogenic and Anticancer Properties of Bifunctional Ruthenium (II)–p-Cymene Complexes: Influence of Pendant Perfluorous Chains. Mol. Pharmaceutics, 12, 3089–3096, 2015. 143. Lai, H., Zhao, Z., Li, L., Zheng, W., Chen, T., Antiangiogenic ruthenium (II) benzimidazole complexes, structure-based activation of distinct signaling pathways. Metallomics, 7, 439–447, 2015. 144. Nowak-Sliwinska, P., van Beijnum, J.R., Casini, A., Nazarov, A.A., Wagnieres, G., van den Bergh, H., Dyson, P.J., Griffioen, A.W., Organometallic ruthenium (II) arene compounds with antiangiogenic activity. J. Med. Chem., 54, 3895–3902, 2011. 145. Sun, D., Liu, Y., Yu, Q., Qin, X., Yang, L., Zhou, Y., Chen, L., Liu, J., Inhibition of tumor growth and vasculature and fluorescence imaging using functionalized ruthenium-thiol protected selenium nanoparticles. Biomaterials, 35, 1572–1583, 2014. 146. Castellarin, A., Zorzet, S., Bergamo, A., Sava, G., Pharmacological Activities of Ruthenium Complexes Related to Their NO Scavenging Properties. Int. J. Mol. Sci., 17, 1254, 2016. 147. Baharara, J., Namvar, F., Ramezani, T., Hosseini, N., Mohamad, R., Green synthesis of silver nanoparticles using Achillea biebersteinii flower extract and its anti-angiogenic properties in the rat aortic ring model. Molecules, 19, 4624–4634, 2014. 148. Gurunathan, S., Lee, K.-J., Kalishwaralal, K., Sheikpranbabu, S., Vaidyanathan, R., Eom, S.H., Antiangiogenic properties of silver nanoparticles. Biomaterials, 30, 6341–6350, 2009. 149. Kalishwaralal, K., Banumathi, E., Pandian, S.R.K., Deepak, V., Muniyandi, J., Eom, S.H., Gurunathan, S., Silver nanoparticles inhibit VEGF induced cell proliferation and migration in bovine retinal endothelial cells. Colloids Surf. B: Biointerfaces, 73, 51–57, 2009. 150. Satapathy, S.R., Siddharth, S., Das, D., Nayak, A., Kundu, C.N., Enhancement of cytotoxicity and inhibition of angiogenesis in oral cancer stem cells by a hybrid nanoparticle of bioactive quinacrine and silver: Implication of base excision repair cascade. Mol. Pharmaceutics, 12, 4011–4025, 2015.

Metals and Metallodrugs in Angiogenesis  317 151. Kang, K., Lim, D.-H., Choi, I.-H., Kang, T., Lee, K., Moon, E.-Y., Yang, Y., Lee, M.-S., Lim, J.-S., Vascular tube formation and angiogenesis induced by polyvinylpyrrolidone-coated silver nanoparticles. Toxicol. Lett., 205, 227– 234, 2011. 152. Ikeda, N.E.A., Novak, E.M., Maria, D.A., Velosa, A.S., Pereira, R.M.S., Synthesis, characterization and biological evaluation of Rutin–zinc (II) flavonoid-metal complex. Chem.-Biol. Interact., 239, 184–191, 2015. 153. Nethi, S.K., Rico-Oller, B., Rodríguez-Diéguez, A., Gómez-Ruiz, S., Patra, C.R., Design, synthesis and characterization of doped-titanium oxide nanomaterials with environmental and angiogenic applications. Sci. Total Environ., 599, 1263–1274, 2017. 154. Freyre-Fonseca, V., Medina-Reyes, E.I., Téllez-Medina, D.I., Paniagua Contreras, G.L., Monroy-Pérez, E., Vaca-Paniagua, F., Delgado-Buenrostro, N.L., Flores-Flores, J.O., López-Villegas, E.O., Gutiérrez-López, G.F., Influence of shape and dispersion media of titanium dioxide nanostructures on microvessel network and ossification. Colloids Surf. B: Biointerfaces, 162, 193–201, 2018. 155. Liu, F., Wang, B., Li, L., Dong, F., Chen, X., Li, Y., Dong, X., Wada, Y., Kapron, C., Liu, J., Low-dose cadmium upregulates VEGF expression in lung adenocarcinoma cells. Int. J. Environ. Res. Public Health, 12, 10508–10521, 2015. 156. Gheorghescu, A.K., Tywoniuk, B., Duess, J., Buchete, N.-V., Thompson, J., Exposure of chick embryos to cadmium changes the extra-embryonic vascular branching pattern and alters expression of VEGF-A and VEGF-R2. Toxicol. Appl. Pharmacol., 289, 79–88, 2015. 157. Nooris, M., Aparna, D., Radha, S., Synthesis and characterization of MFe 2 O 4 (M= Co, Ni, Mn) magnetic nanoparticles for modulation of angiogenesis in chick chorioallantoic membrane (CAM). Eur. Biophys. J., 45, 139–148, 2016. 158. Zec, M., Srdic-Rajic, T., Konic-Ristic, A., Todorovic, T., Andjelkovic, K., Filipovic-Ljeskovic, I., Radulovic, S., Anti-metastatic and anti-angiogenic properties of potential new anti-cancer drugs based on metal complexes of selenosemicarbazones. Anti-Cancer Agents Med. Chem. (Formerly Current Medicinal Chemistry-Anti-Cancer Agents), 12, 1071–1080, 2012. 159. Fayad-Kobeissi, S., Ratovonantenaina, J., Dabiré, H., Wilson, J.L., Rodriguez, A.M., Berdeaux, A., Dubois-Randé, J.-L., Mann, B.E., Motterlini, R., Foresti, R., Vascular and angiogenic activities of CORM-401, an oxidant-sensitive CO-releasing molecule. Biochem. Pharmacol., 102, 64–77, 2016. 160. Min, H., Wang, J., Qi, Y., Zhang, Y., Han, X., Xu, Y., Xu, J., Li, Y., Chen, L., Cheng, K., Biomimetic Metal–Organic Framework Nanoparticles for Cooperative Combination of Antiangiogenesis and Photodynamic Therapy for Enhanced Efficacy. Adv. Mater., 31, 1808200, 2019. 161. Ciccone, V., Monti, M., Monzani, E., Casella, L., Morbidelli, L., The metalnonoate Ni (SalPipNONO) inhibits in vitro tumor growth, invasiveness and angiogenesis. Oncotarget, 9, 13353, 2018.

318  Advances in Metallodrugs 162. Wen, H., Qin, Y., Zhong, W., Li, C., Liu, X., Shen, Y., Trivalent metal ions based on inorganic compounds with in vitro inhibitory activity of matrix metalloproteinase 13. Enzyme Microb. Technol., 92, 9–17, 2016. 163. Wang, J., Chen, D., Li, B., He, J., Duan, D., Shao, D., Nie, M., Fe-MIL-101 exhibits selective cytotoxicity and inhibition of angiogenesis in ovarian cancer cells via downregulation of MMP. Sci. Rep., 6, 26126, 2016. 164. Couto, E.V., Ballin, C.R., Sampaio, C.P.P., Maeda, C.A.S., Ballin, C.H., Dassi, C.S., Yukari Miura, L., Experimental study on the effects of bismuth subgallate on the inflammatory process and angiogenesis of the oral mucosa. Braz. J. Otorhinolaryngol., 82, 17–25, 2016. 165. Parrilha, G.L., Vieira, R.P., Campos, P.P., Silva, G.D.F., Duarte, L.P., Andrade, S.P., Beraldo, H., Coordination of lapachol to bismuth (III) improves its anti-inflammatory and anti-angiogenic activities. Biometals, 25, 55–62, 2012.

11 Metal-Based Cellulose: An Attractive Approach Towards Biomedicine Applications Kulsoom Koser and Athar Adil Hashmi* Bioinorganic Research lab, Department of Chemistry, Jamia Millia Islamia, New Delhi, India

Abstract

Cellulose is a naturally occurring abundant polymer and the most abundant biodegradable material in nature. It had been widely used in medical applications such as wound dressing, tissue engineering, controllable drug delivery system, blood purification, and contemporary research has made advancements in the field of pharmaceuticals industry especially antibacterial. Cellulose had been modified via acid hydrolysis, amidation oxidation, carbamation, etherification, nucleophilic substitution, and other modifications. Incorporation of metals like cadmium, copper, lead, nickel, and zinc can cause both beneficial and deleterious health effects in humans as well as plants. Plants cellulose has long been used in a variety of medical applications ranging from cotton for haemostatic wound dressings to sutures and renal dialysis membrane. Plant-based cellulose, though useful for many applications, is not produced in a pure state. The presence of lignin hemicelluloses and other molecules requires intensive processes to prepare it for medical use. In contrast, bacterial cellulose while it is identical to plant cellulose in chemical structure is produced without contaminant molecules. Different types of metals are found in plants which are responsible for a particular pharmacological activity. Herein, we are reporting characteristic features of cellulose, modifications, and biological applications of cellulose and modified cellulose.

*Corresponding author: [email protected] Shahid-ul-Islam, Athar Adil Hashmi and Salman Ahmad Khan (eds.) Advances in Metallodrugs: Preparation and Applications in Medicinal Chemistry, (319–362) © 2020 Scrivener Publishing LLC

319

320  Advances in Metallodrugs Keywords:  Abundant polymer, biodegradable, renal dialysis, acid hydrolysis, oxidation, amidation, carbamation, etherification

11.1 Introduction Cellulose is a major essential component of the primary cell walls of green plant life many forms of algae and oomycetes. It is the most organic as well as abundant polymer on earth. It is made up of carbon, hydrogen, and oxygen and being considered the most abundant organic compound in the world. This compound gives the cell walls of a plant strength and structure and is the source of dietary fiber. It is the polysaccharides where the repetitive unit (C6H6O5)n  consists of glucose molecules, where n consists. Cellulose is a fibril element of plant cells and can be extracted from a number of natural sources including wood and lignocellulose fibers, cotton, sisal, jute, sugarcane bagasse, curaua, etc. [1]. In our settings, cellulose chains are instructed to form compact micro fibrils that are stabilizes for inter- and intermolecular hydrogen boding [2]. These hydrogen bonding processes make the crystals totally soluble in most organic solvent, and in water, they can also enhance fiber polymer interactions and enhance fiber interactions in polymer composities [3]. Cellulose is needed to break down the cellulose connection β-1, 4 D-glucose. These were not owned by animals, they were made of several kinds of mold and bacteria. Ruminants and termites such as deer, cattle, and horses in their GI tracts host symbiotic cellulose-secreting bacteria that enable them to degrade cellulose into easy glucose [4].

11.2 History of Cellulose Anselme Payen is a French Chemist who in 1839 found cellulose. He separated it from plant matter and its chemical resolution from plants matter and determined its chemical formulations [5]. Hyatt Manufacturing Company used cellulose in 1870 to produce the efficient polymer called celluloid of thermoplastics. But after that, in the production of cellulose rayon (“artificial silk”) in the year 1890, Herman Staudinger determined the composition of polymer cellulose in 1920. The chemically chief compound was synthesized without the use of any enzymes extracted biologically in 1992, by Kobayashi and Shoda [6].

Metal-Based Cellulose  321

11.3 The Properties and Structure of Cellulose Cellulose has no flavor, no smell, and is naturally hydrophilic [7]. It is insoluble in water, and chiral and biodegradable are the majority of organic solvents. It is chemically divided into their glucose units by treating elevated temperature focused mineral acids [8]. It is obtained from Glucose D- units that condense that reduce glycosides connections through β (1–4) [9]. The theme of the linkage contrasts with that of α in starch and glycogen for (1–4)glycosides bonds. Cellulose is a polymer in the straight chain. Nothing like starch does not coil or branch, and the molecule implements an expended and rather inflexible rod as seen conformation helped by the residue of glucose equatorial stretching the various hydroxyl groups on one chain glucose from the hydrogen bond with the same or neighboring chain of oxygen atoms side by side strongly together and making elevated tensile-resistant micro fibrils. That gave malleable strength in the cell wall wherever micro fibrils of cellulose are interconnected into environment of polysaccharides. A huge amount of cellulose characteristics rely on the length of the chain. Cotton and other plants fibers as well as bacterial cellulose have chain lengths ranging from 800 to 10,000 units [10]. Those fragments having precise slight the chain length typically acquired from the breakdown with the cellulose was known as cellodextrins including long-chain cellulose cellodextrins water and organic solvents. A number of unique crystalline cellulose structures were also known to correspond to the position of hydrogen bond between and within stands. The natural cellulose is cellulose lA and close of higher yields are mainly lβ. Cellulose generated in cellulose fibers is irreversible to

β,1 OH

OH O

HO HO

HO

4 glycosidic link OH O

OH

OH

Non-reducing end

Cellobiose Unit

OH

OH

OH

O OH

O HO

O

OH

OH O

OH

Anhydro glucopyranose unit

O OH

Reducing end

Figure 11.1  Representation of a cellulose chain showing the anhydroglucose unit in the chair conformation along with atom numbering the glycosidic link, and both reducing and non-reducing ends of the polymer.

322  Advances in Metallodrugs cellulose II; importance is that cellulose I is metastable and that cellulose II could produce the structure of cellulose III and cellulose IV [10]. It contained 6.17% of carbon, 44.44% of hydrogen and 49.39% oxygen. Cellulose’s chemical formula is (C6H10O5)n as shown in Figure 11.1 where n represents the degree of polymerization as well as number of glucose groups [11, 12]. The cellulose produced and frequently found in a composites with hemicellulose lignin pectin and other substances although bacterial cellulose is quite pure due to the increased chain lengths that has much increased water content and higher tensile strength [13, 14].

11.4 Modification of Cellulose 11.4.1 Acid Hydrolysis In general, nanocrystals themselves can be generated in the hydrolysis platform on cellulose nanocrystals. Sulphuric acid is the most common acid used in cellulose hydrolysis, followed by hydrochloric acid [15]. In a corresponding technique to that used for hydrochloric acid, the Argyropoulos group used hydrochloric acid for cotton fiber hydrolysis, and occasional use of phosphoric acid was also reported [16, 17]. Acid sampling modifies the resulting nanocrystal characteristics. Such isolated sulphoric acid has a large number because of the surface charge inclusion, then a number of sulphate and phosphate groups on nanocrystal surface resulting in the electro-stabilization of this nanocrystal suspension, and acid selection changing the resulting nanocrystal structures. The amount of sulphate groups is based on the moment of hydrolysis and concentration of sulphuric acid, and the resulting nanocrystals result in a much greater surface load density compared to phosphoric acid [18, 19]. These groups have been suggested to be removed by freeing replacement as well as elevated temperature with concentrated sodium hydroxide [20, 21]. While phosphate fragments developed during hydrolysis were produced on the nanocrystals at C2 or C3 using powerful NMR, there is no representation in the nanocrystals [22] location literature. The occurrence of these ionaziable groups was established as a major factor in cellulose nanocrystals’ surface configuration and reactivity [23]. Conduct metric titration was the common technique used to evaluate the amount of alternation during nanocrystal development. A closer estimation of the results highlighted in the document, however, demonstrates a variety in the proportion of

Metal-Based Cellulose  323 identified sulphate groups singing blended bed resin for two out of three specimens and a resin [24] sulphate measurement. This product could be classified as contamination at an extent not understood by the writers as effective. Using the measurements of 131-10-13 1 nm nanocrystals. It is now possible to determine hydroxyl group number on the surface of nano-crystalline cellulose from 1.57 nmol g (0.51 nmol g1 primary) to 1.82 mol g−1 (0.59 mol g−1 general election) for this set of data as reported by Aolbitol et al. [25]. For this hydrolysis technique, this sulphate content identified for cellulose nanocrystals solution is proportional to DS surf ¼ 0.160.19 even in combination with reported earlier sulphate quantities for this hydrolysis method. Another fundamental consideration to be developed in crystal surface cellulose modifying in the existence of nanocrystal surface contaminants emerges from cotton nanocrystals processing. Labet and Thielemans concentrate on this aspect when they studied cellulose nanocrystal modification across polymerization ring-opening with ϵ-caprolactone [25]. It was reported that Soxhlet extraction of bleached cotton with euphoric acid hydrolysis ethanol of cellulose nanocrystals removed a considerable amount of species forming the surface of nanocrystals, including xylobiose 1,6-anhydroglucose vanillic acid and 3,4,5-trimethoxyphenol (Figure 11.2) [25]. The heavy metals are found on the nanocrystal surface reaction without this point of soxhlet extraction and may also be partly responsible for the indicated esters of cellulose sulphate. Therefore, soxhlet extraction with ethanol is highly recommended before any surface alteration order to respond. O

HO HO

OH

HO O

O

OH

OH

HO HO

O O

HO 1,6-anhydroglucose

Xylobiose O OH

MeO

OH

MeO

HO OMe Vanilic acid

OMe 3,4,5-trimethoxyphenol

Figure 11.2  Structure of the impurities found on sulfuric acid hydrolyzed bleached cotton nanocrystals as removed by ethanol Soxletextraction [44].

324  Advances in Metallodrugs

11.4.2 Oxidation Cellulose showed either an oxidation reaction instead to carboxylic acid or aldehyde functionally [26]. Nitroxyl-based oxidation to selectively generate carboxylic acids in main current alcohols and oxidation during periods to produce aldehyde from directly adjacent diols are the two most dominant various methods [27, 28]. Cellulose oxidation 2,2,6,6-Tetramethylpiperidinyloxyl radical (TEMPO) is both as a procedure of even more surface modifications and as a bulk method [7, 29]. Oxidation usually directly recently occurred with a catalytic oxidation quantity (TEMPO) such as sodium hypochlorite or sodium chlorite recycling TEMPU (Figure 11.3). Sodium bromide has also frequently been used to try to boost the oxidation rate through hypobromite formation of even more sodium in situ [28, 30]. At pH < 8, the connection between the two between current main and secondary alcohols was not as slowly and selectively prominent as at 9 < pH < 11, where the research findings showed absolutely outstanding primary alcohol selectivity. This is refused to back by numerous feasible mechanisms’ power calculations suggesting a hampered bridging state under alkaline conditions. This is supported by energy calculations of various suggested mechanisms which really demonstrate a disturbing transitional state under alkaline conditions (Figure 11.4) [31]. This inhibited transitional state demonstrates that compound formation for secondary alcohols is considered extremely beneficial for primary

(a) 2 N

–Cl

–OBr

N OH –OCl

+ –Br + 2– OH

O

O

(b)

N+

RCHO/RCOOH

–Br

N O

RCH2OH/RCH2(OH)2

Figure 11.3  Initial formation of oxidant from TEMPO radical and it catalytically of TEMPO oxidation sodium hypochloride in sodium bromide as stoichiometric oxidant [47, 50].

Metal-Based Cellulose  325

+N

O

H O

-O

OH

N

O

N

O

HHR

OH H

H H R

R

–OH

N

N OH

O H OHR

O HO

R

N O–

O H

OH R

Figure 11.4  Mechanism for oxidation of primary alcohols with TEMPO with a cyclic transition state [49].

alcohols, and this preceding pathway has decomposed the main alcohols under fundamental circumstances [31, 32]. TEMPO oxidation continued to be applied to HCl hydrolysis-extracted cellulose nanocrystals to allow damaging nanocrystal load and mark stability of aqueous suspension [32]. The hypochlorite concentration presence on the oxidation material was scrutinized however on the exterior of crystalline cellulose nanocrystals instead with negligible effect loss [33, 34] showed up to 0.5 hypochlorites/ AGU. Therefore, the application of acetamide-TEMPO under similar reaction conditions has resulted in a more uniformly oxidized nanocrystal cellulose surface resulting in higher stability monodisperse suspension [17]. Also used as  straightness of impurities TEMPO oxidation of cellulose nanocrystals straightness of impurities observed on hydrolyzed sulphuric acid bleached nanocrystals of even more cotton as removed by extraction of soxhlet ethanol. Primary mechanism of alcohol oxidation with cyclic transition [31]. Used as a reference for any further surface characteristics to be discussed later [7]. In the event of cellulose, by selective cleavage of surrounding diols, the 2,3-diol releases the glucopyranose ring and creates two functionalities of aldehydes [35]. The cellulose oxidation level was completed by the suspension in the sodium meta periodate  solution of the cellulose sample (NaIO4) for a particular circumstance particular period of moment [36]. The transformation takes place by connecting the diol with the period anion to form an acyclic intermediate that converts readily to dialdehyde with iodate loss (Figure 11.5) [37]. Iodate oxidation has been shown to

326  Advances in Metallodrugs O HO

O

H O

OH O

O H

O

O

HO

O

OH

O

O

OH

H2O –O

OH

_ O

O

O

O O

–O

OH OH O–

OH O–

OH OH

O O

Figure 11.5  Mechanism of periodate oxidation of cyclic 2,3-diols [31].

have a significant impact on cellulose sample crystallinity that is totally amorphous to DSr ¼ 0.80 (observed) corresponding at a reaction speed of more than 150 h [35]. Curiously, TEMPO oxidation has already been coupled with oxidized surfaces [38]. to isolate nanocrystalline cellulose. Periodate oxidation was performed for 36 h only, and AFM product image shows nanocrystal morphology but no mention of product crystallinity was presented due to the possibility of crystallinity destruction as illustrated above [36].

11.4.3 Esterification Sulfation as well as phosphorylation are therefore signs of the esterification solution that take place throughout the hydrolysis procedure. The only other instances of in situ functionalization in the hydrolysis literature were the manufacture of acetylated and butylated cellulose nanocrystals using blended acetic acid and hydrochloric acid [37, 38] by Fisher esterification. Further, the authors proposed that the methodology facilitates with almost comprehensive transformation of hydroxyl surface to esters with comparative FTIR order to determine esterification concentration. Considering the surface cellulose nanocrystals, there have been important indications that there might be some distortion to the sample’s crystallinity leading to the reduced chance of oxygen bonding with a great extent of substitute on the surface. However, the authors assert that the degree of substitution at 1.5 on the surface of nanocrystals could lead to significant degradation of interchain hydrogen bonds [6]. Taking surface cellulose nanocrystals into consideration, there would be significant signs that there will be some distortion of the sample’s crystallinity compared to a reduced probability of oxygen bonding with a considerable extent of surface modification.

Metal-Based Cellulose  327 In polylactide nanocomposites, nanocrystals of acetylated cellulose extracted with (DSaurf ¼ 0.9–1.05) have been integrated and enriched the temperature with heat decomposition and kinetics of composite material crystallization [37]. Those who use acetic anhydride in pyridine [39] can also perform surface acetylation of cellulose nanocrystals. In this response, pyridine not only performs as a solvent but it’s also about produces an  intermediate reactive driving response future (Figure) though 4-dimethylamino in this position, pyridine (DMAP) has been much more effective [40, 41]. Sassi and Chanzy provided this research with stractural elements of cellulose acetylation spending an acetic anhydride and acetic acid consuming toulene toward avoid cellulose acetate swelling and disintegration. Sassi and Chanzy presented this experiment with structural elements of cellulose acetylation that have used a mixture of acetate acid and acetate anhydride and the use of the toluene to avoid enlargement and dissolution of cellulose acetate. Starting to add a non-swelling solvent with the cellulose acetate shell in a core of unmodified cellulose [42]. This hypothesis could be considered to extend to other heterogeneous esterification of cellulose fibers and nanocrystals of cellulose. The use of both these acid anhydrides in the preparation of cellulose nanocrystals has not been well-founded although the earliest example is the use of alkenyl succinic anhydride (ASA) emulsions in water to extend CNXLs to non-polar media suitable particles. Yuan et al. [43] showed that long-chain alkenyl succinic anhydrid has been used to impart hydrophobic character on cellulose nanocrystals with inadequate reagent extremes leading to DSr ¼ 0.016 which is correlated with DSauf ¼ 0.80 using the nanocrystal measurements cited in paper [43]. The resulting D Saurf cellulose nanocrystals exchanged by benzyl and amethoxy benzyl  pyridinium have been 0.40 and 1.18, respectively [44]. Despite the prominent rate of alteration that has been suggested to modification of secondary hydroxyl groups on the cellulose crystallinity, peripheral was recorded only signifying the addition of amorphous grafts. Transesterification has also been used to change nanocrystals of cellulose. These grafting could be used as follows. By ring-opening polymerization with poly 3-caprolactone [45] but also with acid catalysts, the approach improves cellulose nanocrystals. Due to such a modification, which produces long chains from the cellulose surface and sometimes papers, the composite production and testing of the degree of replacement in these reactions have not been reproved and can not be determined immediately (Figures 11.6 and 11.7) [46, 47]. More lately, transesterification of vinyl acetate in DMF53 has supported out acetylation. The FTIR and 13C CP-MAS NMR technique has been confirmed successful.

328  Advances in Metallodrugs O R

O X

R

N

R O

X= O R

O

X N

O R

S

O

–X

N

–Br –

Cl

O–

Figure 11.6  The mechanism of activation of anhydrides by reaction with pyridine to form a highly reactive actyl pyridinium intermediate. Also applicable to acyl halides and other nitrogen bases [26].

R

H+ N

Br N +

R

N

Br-

N O

O

H

O

R=H, Me

R

R –Br

O

O

Ts

Cel

O O

N

HO

O

–O

OTs

N+ −Br

S

Cl

N+

R

O+

Cel

H N

O

O Cel

Figure 11.7  One post esterification cationization of cellulose nanocrystals as performed by Jasmani et al. [53].

There was no comprehensive quantification. Powder X-ray diffraction was being used to study acetylation time vs. crystallinity and it was uncovered that cellulose nanocrystals structure was significantly compressed after only one extra minute of acetylation (Figure 11.8) [48]. The most prevalent phenomenon to esterify cellulose nanocrystals is the use of acid halide reagents [48–56]. Bromoisobutyryl bromide (BiB) has been thoroughly used to transform cellulose nanocrystals to produce them appropriate for initiating

Metal-Based Cellulose  329 I

C2H5 O

C4H9

Sn O O

O

RO-Sn-OR +2 C2H5 O C4H9

+2ROH

OH

C2H5

II

O

OR O O SnOR

OR Sn OR O

OR O

O Sn RO O

OR Sn

O

OR

Figure 11.8  The mechanism of transesterfication of ϵ caprolactone catalyzed by tin-II 2-ethylhexonate [63]. H

R HO

O

H

O+

R OH

O

O

O H HO

O

O+

O

R

OH H+

R

+ OH O H

Figure 11.9  The mechanism of acid catalysed transesterfication using 3-caprolactone as an example [26].

radical polymerization of the atom transfer (ATRP) from the nanocrystal surface [49, 57]. There are several differences between the published processes. The Hailiang Zang group applied a mild excess of bromoisobutyryl bromide (21mmol g-cellulose) to cellulose nanocrystals suspended in tetrahydrofuran for 24 h with DMAP and triethylamine in a proportion of 1.15:1:1. It already led in a low DSaurf ¼ 0.06. at room temperature of Subsequently, the altered nanocrystals were used for ATRP to procure. Polystyrene (PS) and nanocrystals (PMMAZO) [4-(4-methoxyphenylazo)] hexyl methacrylate (PMMAZO) [51]. The initial social circumstances of 210 mmol g−1 BiB 230 mmol g−1 Et3N in DMF at 7.0 –C led to 24 h in 0.47. DS was realized to be proportional to the temperature of the observation moment and even the concentration of BiB. The lowest possible modification acquired by using 420 mmol g−1 BiB at 70C was DSaurf ¼ 1.00. Most importantly, it

330  Advances in Metallodrugs was expressed that the CNXL structure was reconstructed by using X-ray diffraction (XRD) [51] powder. In good enough condition to completely change CNXLs with BiB, Majoinen et al. used chemical condensation deposition (CVD). This method contributed in 5% Br closest approximation to D Saurf ¼ 1.42 [55]. The organization says a rise of 58 five percent in the alteration from Morandi’s methodology but probably accounts for the differentiation in nanocrystals size used. The application of lower nanocrystals for rearrangement exposures a slightly higher surface area, and therefore, the substantial increase is 42% [55]. The group suggests that the fast rise in the modification has been due to superior dispersion in DMF after preliminary modification with BiB by CVD. This 42% increase is crucial; however,  it came from the cost of a three-fold enhancement in reaction time and an additional step in exposure [51]. It should also be noted that sample adulteration with pyridine hydrobromide is a byproduct if the esterification can not be prevented as nitrogen content is not described in the publication, and the solubility of pyridine hydrobromide in dichloromethane (DCM) used to reconstitute the product is less than 3 mol levels at room temperature [58] of a swelling agent. As the formation of cellulose tripalmitate outside, the crystals fades at 105oC. Therefore, the liquid cellulose tripalmitate formed at the same reaction temperature gets completely dissolved. The esterification gas-phase performs as an antisolvent to retain the previous particle morphology for the cellulose tripalmitate [52]. The scientists indicate that this is speeding up path towards compatibilized cellulose nanocrystals for use in nanocomposites but did not minimize the impact of such significant transformation on the resulting nanocrystals’ mechanical characteristics. Also, under more traditional responding requirements, fatty acid modification of cellulose nanocrystals had been executed. Menezes and Ramie et al. [53] modified nanocrystals of cellulose with chloride fatty acid remained. Their crystallinity in accordance with XRD but the grafting features correspond to D Saurf > 3 [53] according to the elemental analysis indicated in the document. Their crystallinity according to XRD but the characteristics of grafting according to the elemental analysis specified in the paper correspond to D Saurf >3 [53]. This demonstrates either that the nanocrystals have already been altered beyond the crystal surface or that the purity of the product is uncertain. Otherwise, cellulose thermal decomposition had as an easy alternative in the further conversion to cellulose nanocrystals. The Argyropolous cluster used chloride and pyridine p-toluenesulfonyle for two days at room temperature to carry out surface tosylation of cellulose nanocrystals [59]. The result IR spectroscopy was verified, though there was no further theorizing classification so the amount of tosylate and chlorine alteration [60]

Metal-Based Cellulose  331 could not be ascertained. Azidation further encouraged the product to perform click the nanocrystal surface chemistry (Figure 11.9). While coupling agents have been used to esterify Cellulose is prevalent for cellulose nanocrystals in the literature, there are very few explanations. A scientist named Nielsen et al. [61] who used N, No-diisopropyl carbodiimide (DIC) as well as 4-dimethyl aminopyridine (DMAP) to production of methacrylate cellulose [61] first reported it for cellulose nanocrystals in 2010. The report did not acknowledge the esterification accomplishment. The quantity of alteration was recorded as DSaurf ¼ 0.1 after a further reaction, but the incomplete modification of methacrylate groups is noted [61]. Edwards et al. attempted to esterify CNXLs with DIC and hydroxy benzotriazole. However, there is no proof of esterification in the spectroscopic data described in the document and the response was permitted for only 1 h, [62] where full DIC conversion involves 2–3 days for heterogeneous esterification at room temperature [63].

11.4.4 Amidation TEPMO oxidized cellulose nanocrystals are the most prevalent use of cellulose nanocrystal coupling agents as a starting material for cellulose nanocrystal amidation reasons. The method by forming the N-hydroxy­ succinimidly ester monitored by a primary amine response to manufacture form the amide product (Figure) invalidates the activation of carboxylic acid molecules on CNXLs. Araki et al. [64] used this method for the very first moment using a combination of 1-Ethyl-(3-dimethyl aminopropyl) carbodiimide and N-hydroxysuccinimide to amide ultimately aminated poly-(ethylene glycol) oxidized CNXLs with DSaurf ¼ 0.12 [64]. The response stands followed out particularly on the C6 position of cellulose (polysaccharide) owing to the performed oxidation region prior toward the amidation but not all carboxy groups on the CNXLs have been modified. The same technique has been applied later by Filpponen et al. [16] to DSaurf ¼ 0.17 and DSaurf ¼ 0.10, respectively, for amidst oxidized CNXLs with propargyl amine and 11-azido-3,6,9-trioxaundecan-1-amine. With this new technique for Jeffamine (aminetermined polyethers) grafting with a recorded DSaurf ¼ 0.1 [65], Azzam et al. widely reported the currently the highest grafting density. Although the writer conducts TEM analyses of their different samples, there is no mention of the fact of accurate nanocrystal physical dimensions and the authors have chosen literature values intended for the averagenanocrystal dimensions produced through the environmental circumstances used for hydrolysis (crosssection of 26 × 6 nm) [65]. The values are incredibly difficult when you

332  Advances in Metallodrugs attempt to assess the amount of modification on the nanocrystal surface. Achieving a nanocrystal size of 266 nm is actually feasible with the actually original mentioned degree of oxidation (DO) of 0.2 before amidation, as the main hydroxyl groups on the nanocrystal surface are not sufficient to  cause. As the DO remains assessed by titration directly and immediately, the biggest problem must be that the size of much of nanocrystal was presumed. Assuming that for the widely quoted DO of 0.2 a nanocrystal cross-section of 26 × 6 nm results in a DSaurf ¼ 0.5, signifying that completely major economic hydroxyl groups remain oxidized. By means of these minor modifications, DSaurf ¼ 0.28 is almost twice the degree of amidation control for this immediate reaction. A comparable technique was used by Harrison et al. using diisopropyl carbodiimide (DIC) with polymer grafting NHS toward oxidized CNXLs by intimidation DMF [66]. The resulting density of grafting had been comparable in appearance to the preceding research in DSaurf ¼ 0.1. Fujisawa et al. [67] intended to use DIC without NHS between several CNXLs in DMXLs in DMF, but led for the development of N-acylureas advanced O-acylisourea (Figures 11.10 and 11.11) [67] arrangements. This was mentioned before for responses performed with DIC in DMF on Wang resin (solid-state benzyl alcohol) and the withdrawal was not investigated by the treatment in DCM through designed for cellulose nanocrystals. R' O R"

O

R' N

C

N

HN R'

H R"

O

O O

R'

R'

H N

C

N

R'

R"

O

N H

O

N

O R'

O R

R"

NH

O

O

NH

O

N O

R'

NH O

N

N R

H

H

Figure 11.10  Mechanism of amidation using carbodiimides to form an N-hydroxysuccinimidyl ester intermediate [67].

R'

Metal-Based Cellulose  333 R' O +R

HN

O O R'

N

N+ H

R"

NH

O

R

R'

Figure 11.11  Mechanism of formation of N-acylureas through rearrangement of O-acylisoureas [82].

11.4.5 Carbamiation The procedure for the of isocyanates to change the nanocrystals of cellulose could be categorized into dual categories approximate. However, toluene-2,4-diisocyanate (TDI) is extended to assign functional polymers or other molecules, and non-polar isocyanates are being used to change the nanocrystal surface qualities (Figure 11.11). Polycaprolactone (PCL) grafting with TDI to cellulose nanocrystals was the first isocyanate use, reported to modify cellulose nanocrystals. The solution was performed in triethylamine toluene as a catalyst (Figures 11.12 and 11.13) and a reaction of 7 days at 90°C [68]. A technique earlier used by a starch with phenyl isocyanate at one end responded with TDI in 1:1 stoichiometry to create a mono isocyanate that was subsequently combined with cellulose to avoid the interconnection of nanocrystals [69, 70]. Cellulose crystallities [70, 71] surface reaction has been restricted through the use of non-swelling toluene. Zoppe et al. immediately illustrated the same response with a shorter 24-h timeframe in 2009, and although FTIR grafting has not been compatible with either paper percentage. During modification [72], the degree of suitable replacement occurred. Siquiera et al. recovered the first quantity of surface adjustment analyzed using isocyanates from cellulose nanocrystals in 209, which changes cellulose based on sisal with n-octadecyl isocyanate without any catalyst as well for DSaurf ¼ 0.07. This gives gives DSaurf ¼ 0.11–1.2. using the paper (5–1.5 nm) documented nanocrystal average diameter. A subsequent paper by the same authors further analyzed the emotional response and discovered that the reaction modified only 3.7% of the available surface hydroxyls. The authors chose a 5-nm cross-section to determine the size of the nanocrystals but used Gardner and Blackwell’s crystallographic data in 1947 to shut down the number of chains in a crystal. To determine the amount of groups of surface fatty acids. In 2002, information on the

334  Advances in Metallodrugs O

R=Alkyal, Aryal

C

R

N

N C O

N

C

O

Figure 11.12  Isocyanates used in the modification of cellulose nanocrystals.

spectroscopy of particle accelerator X-ray and proton fiber replaced the crystallographic information which showed nearly half of the information given in 1974 [73]. This indicates that the editors’ calculation that 63% of cahin is on the surface in a nanocrystal is mistaken 45% using information from Nishiyama et al. [52] and that 5% of hydroxyls have actually been modified on a 5 nm cross-section crystal layer. Discounting the carboxylic C3 upgrade to 7.8%. The implementation of their alteration is therefore to make it more effective than mentioned now in the article. In additional previous effort to alter cellulose nanocrystals hydrophobically using isocyanates concerned transplantation of phenyl isocyanate-capped caster oil cellulose nanocrystals using TDS [74]. The scientists perform + +

NR3 O

Cel

Cel

H

O C

O

R

N H

H NR3

N R

O

O Cel O NR3

R NH

Cel

R NH

O H

+NH3

Figure 11.13  Mechanism of reaction between isocynates and cellulose (and other alcohols) as catalysed by tertiary amines [87].

Metal-Based Cellulose  335 the modification using different triethyl amine catalyzed feedback circumstances and achieve a full collection of analytical methods. Using elemental analysis, the proportion of alternation was reviewed and determined by comparing the product to be 21% mass change in carbon ontent [74]. Where as, the product seems to have a considerable discrepancy a particular deformation level between carbon and nitrogen content. Whereas that may also be elucidated by Shang et al. [74]. However, the implication is that it is a castor oil combination of triricinoleoylglycerol rather than a pure. Not necessary to take this into consideration in your calculation. The usage of castor oil and other mixtures to improve cellulose sometimes doesn’t allow the surface alternation of cellulose nanocrystals to be effectively assured owing to the imprecise nature of the alternation and this makes it very difficult to estimate the proportion of reaction achievement. TDI was used in the compound improvements of the cellulose nanocrystals. Morandi et al. used TDI to concentrate a photo cellular ATRP initiator in and out of cellulose nanocrystals to encourage the development of polymer brushes which can be easily replaced from the underlying nanocrystals for further analysis [75]. In this ongoing research, instead of dibutyltin dilaurate triethylamine (DBTDL) remained established as an isocyanate catalyst conversions and the interactions have been conducted. At the same or lower temperature 40 μC whereas the experiment remained efficacious (as evaluated some discrepancies are observed in the FTIR, XPS, and elemental analysis) in the performance of the fundamental examination. DSaurf ¼ 0.06 calculated in the observed amount of alternation on the principle of the bromine elemental analysis [75]. Even though this remains applicable outstanding to the possibility of the bromine atom for the next modification stage identified, it would only consequence in 0.34% nitrogen in the product rather than 1.3% in the report. An optional  explanation is an incomplete response to cellulose response between photocleavage graft and TDI. Even before cellulose reaction, there is no purification, and this could result in TDI modification which has not functionalized with the bromine of the photo cellular initiator. The recent modification using an isocyanate has been that hydrogen bonding ideas were attached to cellulose nanocrystals in light-healable nanocomposites. Biyani et al. [76] created pyrimidine- hydrogen-bonding symbolism based on such a group of isocyanates. It has been the case connected with DBTDL as a high temperature catalyst of up to 100°C by the response in DMF to cellulose nanocrystals. Using the 25–26 nm diameter reported by the paper [76], the publishers fail to qualify the modification using visible UV spectroscopy and suggest (DSaurf ¼ 0.18) corresponding to total surface modification (DSaurf ¼ 1.5). Biyani et al. [76] do not sumptuous on their DS technique of intention

336  Advances in Metallodrugs S C N

Et Et

N

Cl–

Et N

O

Et

COOH COOH

HO

O N

C

S

Figure 11.14  Fluorescenic isothiocyanate (FITC) and rhodamine B isothiocyanate (RBITC) used in the modification of cellulose nanocrystals.

but closer inspection of their observational information leads to the calculation by UV-visible spectroscopy of DSaurf ¼ 0.59–0.96 intention but closer inspection for their observational data leads to DSaurf ¼ 0.59–0.96 calculation using spectroscopy with UV visibility. Neither the editors’ values nor those discussed here, however, agree with the outcomes of either the elementary assessment of the alter cellulose published in it [76]. A comparable group of chemical isothiocyanates (Figure 11.14) using fluorescent isothiocyanate (FITC) and rhodamine B isothiocyanate (RBITC), Nielsen et al. [61] responded to cellulose nanocrystals in 0.1 M sodium hydroxide at an average temperature for 72 h. The resulting nanocrystals had a 2.8 μ mol g−1 modification to try to reduce the efficiency of this amount of modification that was sufficient for the assisted application [61].

11.4.6 Etherification Cellulose nanocrystals etherification might be the second most omnipresent method that have been discussed in the literature. The most prevalent etherification of cellulose nanocrystal appears to be the utilization glycidyl trimethyl ammonium sulphate (GTMAC) or derivatives to cationized the surface of cellulose [77, 78]. C Glycidyltrimethyl potassium, a suspension of cellulose nanocrystals, is added to carbonate in 1.75 M solution of sodium hydroxide and heated for a few hours to influence modification [78]. The resulting nanocrystal cellulose has DSaurf ¼ 0.4 for timber-borne nanocrystals connecting to for timber-borne nanocrystals corresponding to

Metal-Based Cellulose  337 DSaurf ¼ 0.1 surface features. Resulting CNXLs have already been hydrolyzed and the distribution of that same replacement has already been determined using approx. NMR. 1:1 O2 and O6 preference with substantially less O3 [78] substitute. The replacement pattern is consistent to the one mentioned in cellulose fiber etherification demonstrating relative reactivity as nucleophilic of cellulose hydroxyl groups. Unfortunately, GTMAC modification solution hydroxide suffers from GTMAC hydrolysis issues due to the sodium hydroxide found in the response. The product’s combination and characterization is further complex as well as different reorganizations in case of high surplus reagent (Figure 11.15) [77]. In order to minimize GTMAC loss through hydrolysis, distinct modification requirements were used. Placed with sodium hydroxide powder, the CNXLs were then put into a small amount followed by DMSO/water several hours heating and before sonification by addition to GTMAC. The resulting nanocrystals on nanocrystals of cellulose had DSaurf ¼ 0.35 much greater than reported prior  shifts such. Using cellulose quoted average dimension (55 nm) as provided by Zaman et al. [77]. This provides D Saurf ¼ 0.78 illustration that demonstrates that morphology like a whisker remains conserved but there’s no XRD information has produced toward evaluation whether such widespread alternation has caused the loss of crystallinity and associated structural integrity or a change in polymorph to cellulose III [77]. Another epoxy regent used epichlorohydrin [79, 80] incomparable  reaction conditions to change cellulose nanocrystals. The first to report this shift are Dong and Roman while using epichlorohydrin to add fluorophores for bioimaging to cellulose nanocrystals, first reacted with such as epichlorohydrin in sodium hydroxide solution followed by another ammonium hydroxide conversion to generate a prominent amine-finished surface. This was a contribution of hydroxide to manufacture a prominent surface finally completed with amines. This was decided to respond to a fluorescent isothiocyanate (FTIC) surface alternation of 0.03 m mol−1 or DSaurf ¼ 0.0188. In this employment, UV-Vis spectroscopy has been used to evaluate the surface modification and there is no verification of the additional acquisition of the original response. In addition, direct interaction between FTIC and unmodified nanocrystals of even more cellulose also results in grafting without epichlorohydrin results. Epichlorohydrin has been used more commonly to attach on the surface of even more cellulose (polysacharides) nanocrystals, b-cyclodextrin [79]. The nanocrystals have been stopped in 2.5 M NaOH through b-cyclodextrin before the accumulation to going of epichlorohydrin [79]. Gravimetric evaluation process and photometric titration determined the extent at least of the grafting as 16.9% weight corresponding to DSaurf ¼ 0.13–0.25. From the

338  Advances in Metallodrugs (a)

Cel OH

O

–OH

N+

O +N

Cl– O

(c) – OH O +N

Cl–

Cl–

OH +N +N

Cl–

(b) Cel O –OH O

N Cl– Cl– N+

HO

OH OH

Figure 11.15  Mechanism of reaction between epoxides and cellulose nanocrystals (a) showing multiple substitution (b) and hydrolysis of the starting material [79].

NMR information gathered from the solid state now in this document, the situation is genuinely unsure whether the circumstances of sodium hydroxide have not been reported an adverse effects on the crystallinity of cellulose nanocrystals or triggers cellulose assimilation exposed to conflicting cellulose signals and cyclodextrin (C6 has been allocated to cyclodextrin at 64 ppm) [79]. Kloser as well as Gray [80] also used epoxide to change poly (ethylene oxide) PEO chains instead in cellulose nanocrystals. CNXLs were mixed with solution of sodium hydroxide prior to epoxy resin-finished PEO general reaction resulting in DSaurf ¼ 0.06 [80]. It is uncertain whether the lower amount of alternation so this is because of a reduced position amount of alternation in this situation due to a lower amount of reactants compared to the previous instances or a differential steric effect in the PEO and GTMAC sizes. It was necessary to record an aryal ether of cellulose nanocrystals by Hassan et al. [81] with powdered hydroxide of sodium, the CNXLs are deferred in DMSO and heated before adding 6’2’-terpyridine and 4’-chloro-2,2’ constant heating for many hours impact O-aromatic substitute (Figure 11.16). Due to the measurements in the document, the resulting DSaurf ¼ 0.15 [81]. The given XRD shows a significant peak magnitude rise in the range 2θ = 20• which may possibly be an substantiation of cellulose II [82] formation. In addition, the supplied solid-state NMR spectrums indicate a signal rise of 83 ppm which has been typically calculated as the maximum in C4 amorphous cellulose associated. This may demonstrate that has been noted alternation be situated because integrated regents, now recrystallized cellulose as a replacement for of always a external modification. Finally, alkyl dimethylsilyl chlorides of variable chain lengths [83] had been used to silylate cellulose nanocrystals with

Metal-Based Cellulose  339 imidazole in toluene. Now, it have shown that the silylation to approach a plateau with DS saurf ≤ 1 develops quickly in the first few hours. If the reaction went for an unreasonably long reaction period or if the silyl chloride concentration dramatically increased, the DSaurf exceeded one and the structure of the cellulose nanocrystal would have been destroyed [83]. Pei et al. [84] used the same reaction conditions. The surface modification level, however, was not investigated in 2010 as the journal’s primary focus was on composite formation and characterization [84].

11.4.7 Nucleophilic Substitution The response of the cellulose hydroxyl group acting as nucleophilic in separate responses has been affected by all the cellulose responses described in the classification so far. Performing cellulose carbon nucleophilic replacement reaction opens up a larger variety of functionalization alternatives than those preceding solutions [85]. A greater reaction wealth was undertaken. The most evident position for nuclear replacement owing to steric considerations has been the primary hydroxyl unit carbon atom C6 through a mechanism of SN2 [86]. Substitution occurs at C2 and C3, however, substitution on dissolved cellulose was taken at secondary position using nucleophiles such as azide or flourides [87]. Research with regard to homogenous azidation of slightly cellulose tosylated (DSaurf ¼ 1.5) showed azide formation on both conspicuous and secondary positioning of the remaining hydroxyl propionylation group was shown previously the primary group replacement [85]. This referred to the hypothesis that replacement passed through a cyclic intermediate at the secondary position that associates with prior cellulose tosylate research that indicated the capability to generate epoxy through warming with base [88]. Therefore, the corresponding epoxide conformation (allo or manno) was not directly conformed [88]. A straight line with 2,3-diaxial substituents (to align the lone pair with the orbital C-O’s S) [89] must be generated. Analysis of 2,3-anhydro cellulose hydrolysis materials illustrates preferential epoxide C3 but cellulose tosyl azidation indications to partial azidation at C2 (allo-epoxide suggestion) [90]. The response with butylamine is the same [91], in spite manno epoxide (hydroxyl asult at C2 over C3) development. This indicates that subsequent C3 substitute product would also favor a mann-exposed C2 nuclear attack resulting in extremely favorable twisted-boat conformation. This raises doubts about the validity of the hypothesis of producing an intermediate type of epoxy in nucleophilic substitution at secondary positions of tosyl cellulose and instead of predominating C2 products. Due to a shorter C1–C4 range resulting in a

340  Advances in Metallodrugs + +

OH Cel

O H Cl

Cel

N

N N

N

N

O

O N

O Cl _

Cl

O

N

Cel

N

N

N N

Cel

N

Cel O N N

N

Figure 11.16  Mechanism of formation of aryal ether of cellulose nanocrystals via nucleophilic aromatic substitution, as performed by Hassan et al. [81].

modification in the AGU conformatiom [92], this transformation is further inhibited in the solid state. This would involve the polymer chain to be shortened promote on the surface of cellulose nanocrystals by breaking many hydrogen bonds, which indicates that nuclear substitute at C2 and C3 is extremely likely. The first nuclear replacement reported on cellulose nanocrystals was chlorination with thionyl chloride (Figures 11.17 and 11.18). Initial OR

OR O

O HO

O Ts

O– OTs

OR

O O

O

o

O O

Base

O

OO Ts

RO

O

OH O O NU

OTs

++ +

+ +

OR O

O

OTs

O O–

O

O

O

O RO

NU

Figure 11.17  Mechanism of formation of 2,3-anhydroglucose, showing two possible boat conformations [86], and the monno-epoxide, which is subsequently opened at C3 [88].

Metal-Based Cellulose  341 O Cel

OH

Cl

N+

O

Cel N

Cl

S

Cl

Cl–

O S

S

-HCl

Cl O

Cel

-SO2, -Py

Cel

Figure 11.18  Mechanism of cellulose chlorination using thionyl chloride with pyridine [93].

nuclear replacement of thionyl chloride with ammonia interaction for nucleophilic substitute of cellulose carbon by generating a healthy group of exits. The reaction was carried out in a combination of toluene and pyridine. Decrease nanocrystal expansion and inhibit suspension. Even though the concentration of alternation with the azide-alkyl cycloaddition were used as a catalyst further nucleophilic cellulose replacement, IR spectroscopy adhered chlorination and azidation of cellulose nanocrystals by tosylation. There was no tosylation rate report. For the width of cellulose nanocrystals, the later azidation concentration acquired in DMF using sodium azide was recorded as DSaurf ¼ 0.015 corresponding to DSaurf ¼ 0.53. This amount of deformation is comparable to all average hydroxyl surface groups showing that, as discussed above, the solution may be restricted to this site.

11.4.8 Further Modification All modification reactions on unmodified nanocrystals of cellulose have been characterized, however, a considerable number of such answers to questions they have been used as precursors for further cellulose alteration. Some of these changes can remain related toward cellulose DS that is shown below. Fluorescent-colored cellulose nanocrystals first identified by Dong et al. [92]. They responded with FITC by changing cellulose nanocrystals instead of using epichlorohydrin and ammonium hydroxide. The first stage of development of alteration was the esterification by various methods of fluorescent labeling of cellulose nanocrystals with methacrylate presented by Nielsen et al. This precursor decided to immediately respond with cysteamine DSaurf ¼ 0. 1 in relation to these amines in conjugation. In relation to these amines in conjugation functional nanocrystals mixed with tetramethylrhodamine succinimidyl ester (TAMRA-SE) and Oregon

342  Advances in Metallodrugs green (OG-SE) 488 succinimidyl ester (Figure 11.4) in the direction of manufacture dual fluorescent symbols with carboxyl fluorescein succinimidyl ester (FAM-SE). The average substitution with the shapes produced by this technique was 15.1 μmol g1 corresponding toward or 12 times of the amine groups becoming attached. The average substitution with the forms produced by this procedure was 15.1 μmol g1 corresponding to DSaurf ¼ 0.01 or 12 times of the amine groups becoming grafted. Hasan et al. have given fluorescent cellulose nanocrystals by interacting terpyridine moieties till after complexing with ruthenium terpyridine perylene complexes [81]. Esterification has been used as a precursor to further modify lysozyme’s connection to nanocrystals instead in cellulose [62]. Cellulose nanocrystals were first completely altered with Fmoc-glycine using DIC as a coupling agent before amidation for lysozyme deprotection and maybe even attachment [62]. The concentration of modification for esterification has already been recorded as DSaurf ¼ 0.09 but lateral proportions for nanocrystals are not given in the publication, indicating the transformation of this value into DS problem. Elazzouzi-Hafraoui et al. [94, 95] used nanocrystal size literature values of DSaurf ¼ 0.38 for their hydrolysis method. The lysozyme attachment would then be evaluated to use nitrogen to protein conversion factor and instead of direct nitrogen concentration measurement in the starting material of lysozyme leading to a figure that can not be directly connected to DS. Amidation has been used as a precursor to the widespread cycloaddition reaction of even more azide-alkyne. Filpponen et al. [17] used EDC to add features and functions of azide and alkyne to oxidize nanocrystals of that same cellulose. DSaurf ¼ 0.10 for all of the azide and DSaurf ¼ 0.17 for the alkyne had the developing nanocrystals. These double forms of synthesized nanocrystals has been combined composed for the formation of nanoplatelet gels carbon-catalyzed cycloaddition azide-alkyne (CuAAC), which would have been compatible with that of the FTIR spectrum loss of the azide band [17]. Also manufactured cellulose nanoplatelet gels to produce azido-deoxy cellulose nanocrystals with DSaurf ¼ 0.073, as well as the Click on the two response components, illustrates a 50% decrease In FTIR azide band intensity indicating a  measurable transformation cellulose functionalized by alkyne 1,2,3-triazole product [17]. Attaching imidazolium and potassium carbonate to nanocrystals of cellulose (Figures 11.19 and 11.20) [93, 96], CuAAC was also completed. Before succeeding nucleophilic replacement without azide anion for the manufacturing of azidideoxycellulose nanocrystals, cellulose nanocrystals have all been chlorinated using thionyl chloride [96]. In this case, the imidazolium salt level of reconfiguration might not have been

Metal-Based Cellulose  343 TAMRA R1/R2

R=1

COO–

NH

O S Cel

O

N

O o

HO

N+

O

cooH

R2-

F

F O

HO

Ο

OG

FAM

Figure 11.19  The finished structure of fluorescently dyed cellulose nanocrystals reported by Nielsen et al. [75]. Cu(I)

Cu (I) NCel N+

Cu (I) R

N Cel +N

N

N

R

R N

N R=

Cu(III)

N

N

N N

Cel

N

Cu(I)

H+

R Cu (I) N N N

Cel

X– N+

Cel

Fe

Figure 11.20  Mechanism of copper catalysed azide-alkyne cycloaddition according to Himo et al. [99] and structure attached to cellulose nanocrystals via this route.

reported for azidation; however,  the simultaneous grafting of the imidazolium cations brought in ion-exchange DSaurf ¼ 0.20 cellulose. The azidation concentration in ferrocene grafting was observed to be DSaurf ¼ 0.56, greater like all primary groups of hydroxyl grafting at the nucleophilic substitution external required. Ferrocene grafting on both of these nanocrystals appeared in an almost maximum estimated modification rate of DSaurf ¼ 0.48 on the surface of cellulose nanocrystals [97] for influential hydroxyl individuals.

344  Advances in Metallodrugs

11.5 Present and Future Medical Applications of Cellulose as Well as Its Components Cellulose is the most harmful biodegradable natural material and was extensively assistance in medical applications, like, as a wound dressing, tissue engineering, controllable drug delivery system, blood purification, etc. Wrapping purpose such as scaffolds in tissue engineering short-term skin substitution hemostatic post-operative adhesion barrier agent and material for hepatocyte culture.

11.5.1 Cellulose Used as Wound Dressing The wound could be stay described as a permanent in any tissue, cut or break generated through injury or chemotherapy. The wound maintenance area has also been widely varied and competitive in the services industry, including usual goods like dry bandage in the direction of complicated hydrogels besides alginate dressing which really contains artificial skin and anti-infective substances used or may be used in wound care. A sticky condition needs to be properly maintained for cost-effective wound healing. A humid environment, as mentioned, works to help to promote healing in several aspects of life. It prevents some more tissue loss from drying and encourages lytic enzyme activity in early wound healing that clear surviving fragments. In the 1900s, it heavily assumed that if they remain dry and uncovered, wounds cure faster. Those materials, however, are very expensive and simply been used for a very short time, so various different biocompatible materials for wound dressing remain going to come up. Recently, twisted management has become more difficult when it comes to new insight into wound healing and a growing need to manage complicated injuries separate the hospital. In order to accomplish cosmetic and functional outcomes, the elementary wound management function and role command express of wound healing. A wide range of available modern dressings has been put on the market, composed of layers mostly with specific characteristics. For wound dressing and and very least significant for connections and connected applications, cellulose-based materials were beneficial. Through preserving the enough the moisture environment in the wound is capable of absorbing structural deficit exudate from cellulosic products. There has been a lengthy and time-consuming convention for using cellulosic brands in medicine which mostly implements particularly interlocked cotton fabrics. Cotton have been appropriated strength to

Metal-Based Cellulose  345 sterilize against steam or heat and provides an excellent appeal to patients and medical executives as a cotton flannel [98]. The following characteristics should be described as an acceptable wound dressing: 1. To maintain the moist environment around the wound 2. Make it possible the distribution of gasses [99] 3. Remove excess exudates to outside surface protects saturation of the dressing 4. Prevents and does not pollute micro-organisms with foreign particles 5. Consider mechanical protection possible 6. Atmospheric temperature control and pH 7. Easy then convenient to eliminate change 8. Decrease the wound’s throbbing 9. Effective cost and reasonable chemically

11.5.2 Dental Applications The bacteriological cellulose generated by the Glucanacetobacter xylinus strain has been tested in human dental tissue regeneration due to its reasonable and mechanical strength of clotting properties. Nano cellulose may have done a lot to help in the recovery with periodontal tissue. Gengifl ex bandage includes a number of double coatings such as an innermost coating of microbial cellulose in which chemical-modified cellulose provides flexibility to the membrane and an outer layer of alkali cellulose [97]. Gore-Tex remains a composite non-resorbable porous membrane composed of urethane and nylon enhanced polytetrafluoroethylene (ePTFE). Now, a complete restoration of an osseous defect around an intra-mobile cylinder implant (IMZ implant) has also been revealed in collaboration with Gengifl ex medication. Gengifl ex therapy’s economic advantages include preserving aesthetic appearance and mouth function. However, it took a small number of surgical interventions. The Novaes and Novaes [98] alternative experiment completed the second restoration of an osseous defect around a TiAl6V4 (IMZ) implant by combining the use of an implant placed in the alveolus of a damaged lower bicuspid and guided tissue regeneration principles. Anjos et al. [100] in 1998 correlated biological performances of two furcation barrier membranes such as ePTFE and cellulose used in mandibular molars.

346  Advances in Metallodrugs

11.5.3 Engineering For tissue engineering as well as other biomedical applications, the plant cellulose similarly retains promise [101]. Different kinds and changes of this cellulose are under comprehensive scientific research and have been tested with distinct kinds of cells for their biocompatibility and bioactivity [101]. The redeveloped cellulose, such as with sanitized cellulose, in which extremely brief from the fibers trees leaves were already chemically completely transformed addicted to fairly lengthy sufficient filaments used in textiles and non-wovens were used to rebuild capillary tube threedimensional tissue [102]. Other than all this, also for respiratory tissue engineering with such as cellulose acetate [103], Non-woven fabrics of cellulose [104], however, the hydrogels [105, 106] based on injectable cellulose have been successfully tested as carriers for in vitro as well as in vivo cartilage regeneration chondrocytes. For their corrosion resistance with bone tissue [107], a viscose cellulose sponge has been investigated, and as cartilage tissue engineering scaffolds though. All though the human organism’s cellulose continues to act as a material which is not degradable or preciseas well as gradually compostable. For example, the time of degradation of intravenously implanted viscose cellulose beads in rats was matter how long than 60 weeks [108]. It is a very slow degradability of cellulose due, for example, lack of enzymes attacking the (β1-4) link in microbial and fungal cells. Its oxidation is an effective method for triggering cellulose degradability. Various methods and numerous oxidizing agents like NaClO2, CCl4, nitrogen oxides, or free nitroxyl radicals [109] can also generate oxidized cellulose. Throughout the plasma enhancement of prison cell morals media fashionable vitro such as happening vivo macrophages, oxidized cellulose has been degradable by hydrolysis or mediated hydrolytic enzymes. Cellulose oxidation brings to the conversion of residues of glucose to components of glucuronic acid-containing groups of –COOH. The presence of only these group has not attenuated the decomposition time of cellulose, and its pH swelling mechanical stability drug loading effectiveness in the water setting and other material comportment instead just towards the polar and negatively charged –COOH groups could remain situated used to functionalize the oxidized cellulose with different biomolecules [110, 111]. From this study, we explored the growth and development of conductivity and phenotypic vascular skeletal maturation muscle cells on two cellulose material kinds (VSMC) that can be used in soft tissue engineering for 2264 Cellulose 2263–2278. The advancements in the implementation of components science and engineering to medicine have been made in the mid-20th century. Even the first vascular graft

Metal-Based Cellulose  347 implants for chemotherapy membrane and total current substitution all have been developed using pre-existing implants which have been initially created in many other purposes [112]. In recent years, however, the focus of development of micro-related materials might have changed to a bottom-up approach in which materials have also been designed and modified to actively produce desired reactions from a specific genetic system using identified chemical and physical indications [113]. Although biomaterials science had been a diverse field now where the need for meticulous materials engineering has been so apparent as in the development of tissue engineering substrates with which living cells must dynamically communicate. There were many different factors adjacent to something which could affect the cell response growing on such a given material. Biocompatibility of a material which really partially depends as to how different proteins adsorb in vivo or the development medium in vitro at the interface between both the material and the blood or tissue fluid. The adsorption free energy that can also be produced in protein adsorption was related to the hydrophobic and hydrophilic balance and was regarded to be the biggest factor which might influence the reaction of cells [114]. All critically significant variables that helped to the general biocompatibility and bioactivity of a specific material have been the addition of mechanical properties, chemical functionality, surface stress topography, as well as surface roughness [115]. The intention of tissue engineering was to use living cells in a multitude of ways to restore damaged tissue. There are usually two substantial tissue engineering methods. In another responsible manner, in a site of injury or tissue loss, a scaffold device with some kind of has been inserted the easiest way. It must be expected that perhaps the patient’s own cells should migrate into and populate the scaffold that decomposes the lost tissue over time. In the second major method, allogeneic or autologous cells are manufactured in vitro on biodegradable and bioactive scaffolds which imitate the extracellular matrix to guide cells in their development as well as development in order to generate 3D tissue-like material for implantation [116]. Many of the other features may directly affect the efficiency of tissue engineering scaffolds. The scaffolds suggested must be up to the task of 3D porous bio resorbing and biocompatible. They must also have mechanical properties that match those of native tissue and degradation structures that match a new extracellular matrix’s rate of synthesis [117]. Many other innumerable efforts have been made to create tissue-­engineering scaffolds for a wide range of different types of tissue and organs. Mentioned for the engineering of collagenous tissue [118], including such self-assembled peptide amphiphile fibers and polyesters, and layer-by-layer images of polyanions and polycations for regenerative medicine in the liver [119] have been

348  Advances in Metallodrugs proposed. Polycaprolactone hydroxyapatite composite nanofibers have been used to produce bone using mesenchyme stem cells [120] and decellularized organic material had been used to generate larynx [121] material.

11.5.4 Controllable Drug Delivery System As a continuous wound dressing and drug delivery system, an  elevated potential as well for bacterial cellulose itself has been clearly demonstrated. BC, however, has heavily invested in dental talks therapy apps such as dental extraction or epithelial cells transplantation whenever carrying both drug-loaded together. Almost all of these applications could benefit from a substance that breaks down under physiological and antibiotic environments. An increasing number of studies investigating BC-based drug systems for dermal applications have also been reported for many years [122] which offers a systematic up-to-date description of positive consequences; the drawback and barriers in BC’s emerging study sector were its lack of enforcement against infection. To completely destroy the limitation of BC in wounds regeneration recent developments such as BC sponges equipped with amoxicillin [123]. Many drug delivery systems based in BC illustrate a biphasic drug release characterized by an original burst and the second stage of slow-release that can occur from within hours to a few days [124]. This delivery system provides an expanded conservation duration of up to one week with strengthened mechanical and antimicrobial properties as well as a high concentration of injury dressing biocompatibility.

11.5.5 Blood Purification Total parenteral nutrition would be a procedure of purifying a semipermeable membrane from whom the kidneys do not generally work with more than just thousands of membrane fibers that are comparatively commonly recognized as hemodialyzers in a single membrane module. Membrane expertise is carry on to develop while waiting for successfully used during the management of such hemodialysis in the treatment of patients acute renal disease and kidney insufficiency (ESRF). ESRF was a situation at which the durable inability of both the kidney might have caused a dramatic reduction in glomerular filtration rate below 5 ml/min [125]. Due to various metabolites from metabolic reactions, renal failure in the body would be improved overtime where a huge number of molecules are called uremic syndrome [126]. Due to various kidney failure, metabolites from metabolic reactions throughout the body will improve overtime where a huge amount of molecules are called uremic syndrome [126]. The classification

Metal-Based Cellulose  349 was based on the weight of both the molecules and the protein binding capacity including such albumin. Uremic poisons below 500 Da are considered as tiny water-soluble molecules, although they may be called the molecules of the center with MW varying beginning 500 Da to around to 15,000 Da. Purely on the basis of the MW that when a uremic toxin may bind to a protein connected to the protein-bound category. The main component of the hemodialysis apparatus was generally a semi-permeable membrane dialyzer were arranged in the center to form neighboring separate compartments for blood as well as dialysate. Urea β2-microglobulin creatinine which really eliminates surplus water and electrolytes such as potassium carbonate and then bicarbonate. Hemodialysis includes water motion which really includes minnor components in a solution through diffusion and ultrafiltration (UF) through semipermeable membranes. The two basic processes engaged in continuing renal replacement therapy are diffusion and UF. Diffusion can be referred to as the motion of solutes from the concentration of high solute to the formation of reduced solute [127]. Dialysate runs counter to the flow of blood throughout maximizing the membrane the application incline of solutes aimed at appropriate diffusion. Small molecules, the same as urea, simply move smoothly into the dialysate fluid along with  the osmotic pressure. Diffusive approval of a substance frequently be contingent on the blood-dialysis liquid deliberation of blood gradient and dialysis the movement rates and membrane features such as diffusion coefficient on the electrical charge of its MW. Removal of alternative was preferably the flow speed is directly proportional of dialysate [128]. Temporarily, UF was a process whereby, as a result of a concentration gradient, solute was transported through some kind of semipermeable membrane by a fluid. That’s what a heading on in the healthy kidney of man. An UF amount depends upon permeability of the membrane and the hydrostatic blood pressure based on the flow of blood. This procedure has been very effective in eliminating fluid along with early-sized particles size which have been thought to cause uremia. Throughout the preceding development of hemodialyzer, cellulose membranes have been used. They have always been created from omnipresent cellobiose as a saccharide. These membranes were curved with respect to morphology, showing a significantly uniform resistance throughout the thickness of both the wall to mass transfer. Besides these membranes, the low average pore size and the prominent hydrophilicity [129, 130] are characterized because of their specific suitability for a diffusion-based operation, long length of popularity was contributing. The structure of the hydrogel makes it easy to obtain a mixture of low wall volume and increased porosity. The whole underprivileged biocompatibility mechanism for examople cuprophan

350  Advances in Metallodrugs originates the proclamation of huge volumes of reactive oxygen species (ROS) by neutrophils that activate its membrane mostly with hemodialysis [131, 132] and also automatically trigger neutropenia [133]. Chronic ROS releases date complication that either actually causes anemia, amyloidosis, significantly accelerated atherosclerosis, and malnutrition [134]. The earlier research revealed that platelet-leukocyte aggregation was caused by different membranes due to activation of shear stress contact or aganist activation. Only these aggregate cells might be automatically trigger atherosclerotic commemorations as well as a real serious experimental physical condition especially involving platelet-neutrophil interactions is closely associated with septic shock pathophysiology and various different organ system failure [133]. Scientists and researchers should expect further use of adsorptive membranes in the future had already recently begun developing new portable hemodialysis electronic devices imitate the natural kidney and its functions to constantly cleanse the toxins in the blood. Once recovering the dialysate vertebral into the hemodialyzer, a regenerating dialysate technology would have been decided to clean the spent dialysate. An adsorptive tissue would also be a flawless contestant to be using sorbent to replacement the current regenerating dialysate technology. However, the membrane must have a huge capacity for adsorption of relatively small and scale molecules.

11.5.6 Wrapping Purpose Paper is manufactured of wood which really varies depending upon forest resources. As a result, immense rainforest areas have been destroyed annually near to fulfill the wood fiber supply. The frame is made from recycled or virgin cellulose fibers as well as and its physicochemical properties are predicated continuously the kind structure of the material used [135]. Paper could be prepared from practically any type of fiber, from old jeans to grass clippings [136]. It is typically used for printing illustrations on or as wrapping material [137]. Recorded that tons of paper might intentionally produced for packaging or wrapping. Millions of trees are dropping every day since the use of forest fiber might have recently increased significantly [138]. Contentious seems to be the controversial discussion regarding logging and opting instead for non-tree fiber. In the viewpoint of the complete lack of standardized supplies for non-wood pulp manufacturing industry and agricultural residues, renewed interest was also appealed [139]. In addition to adding, environmental problems have increased a need to use non-wood pulp as a low-cost raw material of papermaking [140]. The cellulose was now

Metal-Based Cellulose  351 rendered via wood pulp from wood. A number of processes are used in which eliminating the bulk of the noncellulosic matter would be the overall impact. The first and most commonly been using were the sulfite process using a calcium bisulfite and sulfur dioxide solution, the sodium process using sodium hydroxide and hence the sulfate method using  sodium hydroxide and sodium sulfide solution. The sulfite process is now most commonly used for chemical purposes when these pulps are usually ready and contain approximately 88%–90% a-cellulose, but it might be enhanced by alkaline purification and bleaching. Cellulose esters have been used in applications where toughness, hardness, as well as excellent appearance, high gloss, high clarity, and a good variety of colors are essential. Mostly, they are defined as having a “hot feeling”. A significant ecological competitive advantage was that they were produced from a renewable source. Whether excellent electrical insulation properties, heat resistance, weather resistance, chemical resistance, and dimensional stability are important, they are not suitable. Injection molding is the main processing technique for cellulose esters. There is no significant issue both with processing provided measures are taken to prevent overheating and the granules are dry. The ophthalmic instrument handles toys and sporting products, and fancy products such as toothbrushes, combs, and hair slides, along with spectacle frames are the main characteristic applications. Sequins have become an extremely common application for CA which really contains 33% plasticizer. CAB can be used for tabs, steering wheels, goggles, bathroom fittings, car, and consumer durables ornamental trim, hydraulic system traps, as well as pens drawing stencils. In the graphic design, cellulose triacetate film will be used for greeting cards and specialized electrical applications like semi-conductive separators.

11.5.7 Renal Failure Although cellulose movies were primarily used mostly for wrapping purposes, they were also used in the treatment of congestive heart failure

11.6 Conclusion Many of the literature responses so far have focused on compatibilizing cellulose nanocrystals with matrices for composite material formation. All researchers pay full attention to mainly the mechanical characteristics of the resulting compoites than the chemistry that occurs on the surface of the cellulose. Esterification, amidaion, etherification, carbamation, nucleophilic replacement, and clack chemistry have all been recorded as methods

352  Advances in Metallodrugs for modifying the surface of cellulose nanocrystals in responses where DS was reported or anticipated from the literature outcomes. Esterification has resulted in the largest recorded surface DS at 1.5 but generally the nanocrystals’ average alteration is much smaller. From the figure, it has been seen that esterification, carbamation, and nucleophilic substitution lead to the highest average modifications, although etherfication with substitution well beyond the surface of the nanocrystals have beeen reported but without evidence of retention of nanocrystal structure. The reactivity of cellulose as an elecrophile has supposed to be limited to the C6 position due to unfavorable rearrangements required for reaction at C2 and C3, but some reactivity has been reported at these positions without acceptable explainations. The reactivity of C3 combined with the predictable harmful effect of high amounts of modification on the hydrogen bond network means that in cases where DSurf > 1 is reported the stractural integrity of the products should receive particular attention. Quantification of surface alternation on cellulose nanocrystals is a stimulating problem within the field with heavy requirement on elemental analysis as a technique to determine level o modification. Using a substance technique to quantify a surface phenomenon is an fundamentanlly problematic meyhodology. Certain amount have verified that the crystallinity of the products of the modification reactions and comparing with the starting materials. This is least common in the literature that it should be. The increase in variety of modification carried out on cellulose nanocrystals is promising for future applications with in the field but increased analysis of the extent of modification including cross substantiation with different analysis techniques and hazardous consideration of the evidence of each reaction should be a top priority for future publications.

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12 Multifunctional Nanomedicine Nobel Tomar1, Maroof A. Hashmi2 and Athar Adil Hashmi3* Department of Chemistry, J.C. Bose University of Science and Technology, Faridabad, India 2 Department of Biosciences and Bioengineering, Indian Institute of Technology, Bombay, India 3 Bioinorganic Research Laboratory, Department of Chemistry, Jamia Millia Islamia, New Delhi, India 1

Abstract

Nanomedicine is an interdisciplinary field that integrates physics, chemistry, materials science, biology, and pathophysiology principles toward prevention, diagnosis, and treatment of diseases using engineered nanodevices and nanostructures. Nanomaterials and biological structures are approximately of the same size, which allows for unique interactions between biological systems and synthetic materials for analytical, diagnostic, and therapeutic applications. Nanodrugs can work by very specific and well-understood mechanisms, with more useful behavior and less side effects, thereby enhancing patient compliance The most advanced nanomedicines are multifunctional nanomedicines, capable of simultaneously diagnosing and targeting drug to specific molecular targets by incorporating active molecules, targeting ligands, and imaging agents. Nanomedicine can make further effective impact by implementing cooperativity principles in medicine. The main advantage of long-circulating nanocarriers is their accumulation in the tumor via EPR, which leads to increased drug payloads in the tumor. Another attractive strategy involves conjugation of metallodrugs to bio macromolecules, such as antibodies or functional peptides. However, the toxicity issues of the developed nanomedicines must be investigated to prove their safe and efficacious use. Therefore, future design of safer nanomedicines should be based on the detailed and thorough understanding of biological processes. Better synthetic design strategies would strongly benefit from a better understanding of metallodrugs in terms of liberation, absorption, distribution, metabolism, and elimination (LADME) processes, and their behavior in tissue, cells, and specifically at the bio molecular target site. *Corresponding author: [email protected] Shahid-ul-Islam, Athar Adil Hashmi and Salman Ahmad Khan (eds.) Advances in Metallodrugs: Preparation and Applications in Medicinal Chemistry, (363–402) © 2020 Scrivener Publishing LLC

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364  Advances in Metallodrugs Keywords:  Nanomedicine, engineered nanostructures, tumor, antibodies, toxicity, metallodrugs, target cells

12.1 Introduction The use of biotechnology at nano level in medicine is known as nanomedicine. It basically deals with both therapeutics (for specific drug delivery) and diagnostics (rapid monitoring and targeted therapies). Nan­ omedicine and medical nanotechnology are now important components of research and development all over the world. This discipline includes overlapping of multiple branches from materials science and engineering to cell biology. As we know, nanomaterials and biological structures are approximately of the same size, this allows for unique interactions between biological systems and synthetic materials for analytical, diagnostic, and therapeutic applications [1]. Nowadays, medical field is concentrating its approach towards improving human health by understanding the cellular mechanisms in living cells using nanomaterials for early diagnosis by efficient imaging and targeted therapy. Nanomaterials have a high ratio of surface area to volume, which enables high loading of drugs on nanomaterial carriers. Hollow polymeric nanomaterials can be used to carry more than one drug inside the vehicle for their controlled release. Even the size-dependent properties can be used to tune nanomaterials used for imaging techniques such as quantum dots and super paramagnetic NPs. Nanotechnology can play a key role in cancer diagnosis and therapy. This can help in integrating both so that therapy can be designed based on live diagnosing of cells. Traditional chemotherapy in cancer treatment is associated with nonspecific drug delivery system resulting in side effects and toxicity. Thus, nanodevices can be developed which can give the exact location of tumor cells and target specifically them thus saving the patient from the toxic side effects of medicine. Nanomedicine platforms include some of metallic nanoparticles such as gold and silver [2, 3], liposomes (small lipid vesicles) [4–6], dendrimers [7], and carbon structures such as nanodiamonds (NDs) [8]. Nanoparticles have generated substantial interest due to their ability to integrate therapeutic and diagnostic capabilities, treatment modalities, and imaging techniques when varying conjugations are made using diverse classes of drug/imaging compounds, antibodies, peptides, hormones, or ligands. Labeled and non-labeled nanoparticles are already being tested as imaging agents in diagnostic procedures such as nuclear MRI [9]. These nanomedicines can be used effectively in diagnostic imaging with systems such as optical/CT, optical/PET, PET/CT, and

Multifunctional Nanomedicine  365 PET/MRI [10]. Combinations of multiple imaging agents and pharmacologic drugs on a single nanocarrier have been used to monitor drug release, delivery, and therapeutic efficacy [11]. These nanomaterials can also be used for advance applications such as microchip-based drug delivery systems which are incorporating micrometer scale pumps, valves, and flow channels which allow controlled release of drug/s on demand such as DNA microarrays, protein microarrays, and cell chips. These can be synthesized by various different methods such as CVD and electron beam evaporation. This chapter will focus on multifunctional nanodrug delivery and nanodiagnostic imaging systems and illustrate their variability in potential therapeutics and clinical utilization. The safety and biocompatibility of each platform will be discussed, along with the challenges and future directions of developing multifunctional nanomedicines. Nanomedical developments range from nanoparticles for molecular diagnostics, imaging, and therapy to tissue engineering strategies for restoration of biological functions and regeneration of damaged tissues and even organs. Therefore, nanomedicine is comprised of nanodiagnostics, targeted drug delivery, and regenerative medicine as shown in Figure 12.1.

Improved Solubility Drug delivery across BBB and blood cochlear barrier

Drug at desired site of action Objectives to explore nanomedicines

Theranostic applications

Regenerative medicine Controlled release for longer time period

Figure 12.1  Objectives of nanomedicine.

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12.2 Diagnostics and Imaging The nanomaterials used for diagnosis and imaging basically range from 1 to 100 nm range. The usefulness is basically decided by the size and chemical composition. This size range is optimum not to become entrapped in the microstructures of the lungs, capillaries, kidneys, and liver, and thereby cause blockages, while they are large enough that their clearance time in tissues and organs is long in relation to small molecules. Thus, they are useful where we need to enhance imaging long enough to obtain useful diagnostic information and where we want to release therapeutic agents over time [12]. The nanoparticles can easily pass through the infectious cells linings which are dilated due to inflammation thus enhancing the imaging of diseased areas. The nanomaterials used for biological applications should follow certain requirements. Such as they are biodegradable, and degradation should not occur in aggregates, non-toxic, stable in aqueous media, and buffer solutions, should have strong optical activity. Nanoparticles are used for computed tomography (CT), magnetic resonance (MRI), optical coherence tomography (OCT), fluorescent microscopy, infrared (IR), and ultrasound imaging. In case of cardiovascular diseases, imaging of soft tissues is done using nanoparticles as contrasting agents which incorporate high density contrast generating species. Imaging covers a wide spectrum of technologies and applications from structural imaging focused on morphology to imaging with respect to time to understand the physiological processes such as metabolism, or tracing of isotopes of common elements such as potassium, sodium, calcium, and sulphur. X-ray CT is an efficient tool nowadays used for diagnosis of bone pathologies, calcification of tissues, various cardiac, and vascular procedures [13, 14]. During CT acquisition, X-rays are projected at different angles across the body, and a detector collects the transmitted photons. Then, the numerical analysis allows the 3D reconstruction of images representing the attenuation of X-rays by the biological tissues. Conjugated gadolinium (Gd) nanoparticles are used efficiently for CT imaging rather than conventional iodine crystals which have drawbacks of toxicity and rapid clearance rate due to low molecular weight. MRI is a powerful technique to explore both morphological features and functional changes. Like X-rays and CT, this is capable of producing 2D and 3D images but it has advantage over CT as MRI does not use harmful ionizing radiations so any individual can be exposed to multiple scans which is not the case with CT. In particular, SPIONs (Super paramagnetic Iron Oxide Nanoparticles) can be used for enhanced MRI imaging by

Multifunctional Nanomedicine  367 dispersing them in dendritic molecules. The Iron NPs are less than 20 nm size in which all electrons have spin in the same direction, thus resulting in stronger magnetic field than larger size Iron oxide particles. The interactions between spin of water protons and SPIONs cause change in the spin–spin relaxation of water molecules around the nanoparticles and thus reduce the T2 relaxation time and generate a darker image [15]. SPIONs have advantages compared with conventional contrast agent including higher relaxivity, more sensitivity, good biocompatibility, and biodegradability [16]. In one study, patient with prostate cancer was injected with dextran coated SPIONs. It was observed that the circulation time was increased due to coating thus increasing the diagnostic sensitivity. Next generation contrast agents prepared are having core shell structure which are used to image two different techniques simultaneously, thus improving the sensitivity and accuracy of detection. For example, in case of MR/ optical imaging simultaneously, SPION is coated with biocompatible shell like dextran and functionalized by Cy5 or other fluorescence probe which are used for optical imaging [17, 18]. Magnetic resonator nanofabricated coatings can also be used to eliminate the magnetic susceptibility artifacts caused by the stent material enabling MRI imaging of clots and renarrowing of blood vessels inside the stents [19]. Magnetics’ Gastromark was the first product of this class to improve MRI imaging in abdominal structures. Bayer Schering’s Resovist is another example of FDA-approved iron oxide nanoparticles for liver imaging [20]. Gold nanoparticles show increased emission intensity and are known to overcome limitations of conventional contrast agents (organic dyes) such as poor photo stability, low quantum yield, and low in vitro and in vivo stability [21]. Gold nanorods have been shown to produce a uniquely strong optoacoustic (OAT) contrast effect. OAT is used to produce deep tissue images based on ultrasound effects produced by light absorption. OAT has been of interest as a technique to image tumors, based on differential blood content from normal tissue. The optical absorption in gold nanorods is over 1,000 times stronger than that of organic molecules [22]. Nanoparticles can be used to deliver targeted energy to kill pathological cells. With functionalized nanoparticles, radiation can be delivered to a tumor or to individual cells selected by interaction with antibodies or other ligands attached to the nanoparticle using photodynamic therapy. The optical radiation absorbed by quantum resonant nanoparticles is converted into heat in the surrounding microenvironment. This local heat can be exploited for killing cancer cells and for remotely releasing drugs from the nanoparticle [23].

368  Advances in Metallodrugs Fluorescent dye molecules are basically used for bio imaging. But they suffer from various drawbacks such as narrow excitation bands, small stokes effect, broad fluorescence bands, and photobleaching. Recent studies have focused on developing QDs for biological imaging, in near-IR range having size dependent tunable emission wavelength ranging from 700 to 900 nm and are quite stable to photobleaching. Light within this range has its maximum depth of penetration in tissue and the interference of tissue autofluorescence (emission between 400 nm and 600 nm) is also minimal [24, 25]. At present, QDs are considered to be potential candidates in biological applications, ranging from molecular histopathology, disease diagnosis, to biological imaging [26, 27]. The quantum dots are modified at the surface to prevent aggregation and reduce non-specific binding. Quantum dots (less than 10 nm) like CdSe can be used for in vitro research diagnosis which is 10 times stronger than the organic dyes. When QDs are illumed with UV light electron gets excited between energy levels and thus emitting energy in the form of fluorescent light [28]. The QDs absorb beyond the range in which the living tissues show autofluorscence. Thus, NIR fluorescence shown by silver chalogenides, tellurides, and selenides (Ag2S, Ag2Te, and Ag2Se) has a benefit of deeper tissue penetration and the low aqueous solubility of silver allows minimum release of silver ions in environment, thus making it safe for biomedical applications [29–31]. Similarly Carbon QDs have application in bioimaging due to their unique optical properties and low cytotoxicity when compared to semiconductor quantum dots. Red fluorescent CDs were reported to be used by Ge et al. for in vitro and in vivo imaging. They were shown to be retained in higher concentration in the tumor cells due to EPR effect and thus giving a strong fluorescent signal for tumor cells compared to other cells [32]. After metallic QDs, some of the semiconducting π conjugated polymeric nanomaterials (less than 30 nm) which are known to be wide band gap semiconductors can also be used for bio imaging since they possess direct band gap and thus tunable electrical and optical properties (tuning π-π* band gap). A conjugated complex of streptavidin (SA) polymeric dots was prepared and used to label the cancer cell-surface marker HER2 in human breast cancer cells through the specific recognition between biotin and streptavidin [33]. Squaraine-based polymeric dots with large Stokes shifts and narrow-band emissions in the NIR region were synthesized and employed for labeling receptors on the surface of human breast cancer MCF-7 cells [34]. Similarly, CNTs also possess strong NIR absorption which can be used for photoacoustic imaging. Nanomaterials can also be used effectively for enhancing the detection of other biological samples. Such as in the detection of hCG molecule during testing pregnancy, the red colored line which was formed due

Multifunctional Nanomedicine  369 to antigen-antibody interaction was much deeper and brighter in case of using gold nanoparticles than organic chromophores [35, 36]. Similarly, nanowire hybrids can also be used as submicrometer sensors for both intracellular and extracellular environments like ZnO NWs can be used for detecting glucose levels [37] and in pH measurements [38].

12.3 Drug Delivery and Therapy Drug molecules simply diffuse and distribute freely throughout the body, resulting in undesirable side effects and limiting achievement of proper doses required to bring about efficacious responses. The nano-based drug delivery systems are considered efficient drug delivery carriers due to several factors such as high ratio of surface area to volume which permits high loading capacity, high stability, and possibility to incorporate hydrophobic drugs by enhancing absorption into selected tissue due to targeted delivery, improving intracellular penetration, controlled release [39–42], thus overcoming the limitations associated with conventional drug formulations. Several nanodrug delivery systems are developed such as organic, inorganic, and hybrid drug delivery nano-platforms.

12.3.1 Drug Delivery by Organic Nanomaterials 12.3.1.1 Liposomal Drug Delivery Liposomal nanoparticles are one of the ideal choices for drug delivery due to its amphiphilic nature, biocompatibility, biodegradable, enhanced cellular uptake, and low immunogenticity as shown in Figures 12.2 and 12.3. They are basically made up of non-toxic phospholipids and cholesterol having a spherical vesicle system. The liposomes can be easily coated with inert, biocompatible polymers such as PEG which enhances the circulation half-life. The first FDA approved medicine Doxil is a lipid-based PEGylated nanomedicine used for cancer [39]. PEG is a flexible polymer which reduces the adsorption of plasma protein and biofouling of nanoparticles along with increasing the blood circulation half-life [43]. Some other liposomal drugs used in clinical practice today include AmBisome (amphotericin B liposomes), DaunoXome (daunorubicin liposomes), DepoCyt (cytarabine liposomes), and Visudyne (verteporfin liposomes). A lot of strategies are followed nowadays to increase the circulation time of liposomes in the cells, like coating with biocompatible non degradable polymers such as done in case of Doxil, or by attaching the targeting ligands such as antibodies, folate,

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Target cell Normal endothelium Liposome with ligand attached for active targeting

Inflamed endothelium Liposome for passive targeting

TRENDS in Pharmacological Sciences

Figure 12.2  Active and passive targeting of cells for drug targeting using liposomes [34].

and transferrin [44]. A liposomal formulation of vincristine developed by INEX Pharmaceuticals (British Columbia) is Onco TCS for treatment of aggressive non-Hodgkin’s lymphoma. Its clinical trial data (phase I and II) have demonstrated that Onco TCS has higher accumulation in cells and longer blood circulation time, thus showing more sustained drug release profiles than free vincristine. Different types of pH-responsive polymers can be used such as polyalkylacrylic acid, polyphosphazene, polyglycidols, and polymalic acids for making liposomes sensitive to drop in pH [45, 46]. Preferential intracellular drug delivery can also be enhanced by incorporating pH sensitive phosphatidylethanolamine or oleyl alcohol into the liposome membrane, thus increasing their stability in blood during phase transition [47]. So, the mechanism of drug delivery from the liposome depends upon the type of polymer used either by destabilization of lipid bilayer or by fusion of liposomal and endosomal membrane. It has been shown in some cases that positively charged liposomal membrane shows

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Figure 12.3  Benefits of solid lipid nanoparticles in delivery of doxorubicin.

higher binding and internalization by endothelial cells associated with angiogenic tumor vessels than normal cells [48]. New advances have been made by developing cationic liposomes leading to successful delivery of small interfering RNA (siRNA) [49]. So, nowadays, lot of effort is being put to take lipid-based nanodrugs for clinical trials which do not suffer from traditional drawbacks of liposomes such as rapid degradation, instability of nanoparticle, rapid clearance rate from the body by the phagocytic cells of reticuloendothelial system, high cost, and oxidation of phospholipids.

12.3.1.2 Polymeric Drug Delivery The nanomaterial delivery vehicles can also be coated with polymers, such as polyethylene glycol, to increase their half-life in the blood circulation, prevent opsonizing proteins from adhering to the nanomaterial surface, and reduce rapid metabolism and clearance. Moreover, the use of nanomaterials for drug delivery may minimize adverse effects by preventing the nonspecific uptake of therapeutic agents into healthy tissues. The polymeric drug delivery can be done by various agents as polymerosomes and polymeric micelles. The polymeric drug delivery includes both synthetic as well as natural polymers. Synthetic biodegradable polymers Polyglycolide (PGA), polylactic acid (PL), and PLGA are considered as good candidate for drug delivery due to their small size, easily blend with other polymers, nontoxic nature, and stability. The polymeric

372  Advances in Metallodrugs nanoparticles act as carriers for drugs by either getting adsorbed at the surface or encapsulated inside the polymeric nanoparticles. Polymeric micelles are nanoscale core shell structures obtained by self-assembly of amphiphilic block co-polymers. They are small sized than liposomal particles, self-assembled colloidal nanoparticles having hydrophobic core and hydrophilic shell which prolongs the circulation time in blood by protecting from reticuloendothelial system. They self-assemble at particular CMC from a variety of biocompatible polymers such as chitosan, PEG, methyl methacrylate, poly (amido amine) (PAMAM), and others. The polymeric nanoparticles improve the delivery of drugs due to their ability to protect the therapeutics until it reaches target site. The polymerosomes consist of one hydrophobic layer with two hydrophilic polymer faces [45]. They are more stable than liposomes but with smaller vesicle size and thinner membrane. The hydrophobic drug gets entrapped into the micelle core by covalent bonding, thereby increasing the water solubility of the hydrophobic drugs, thus saving the body from the side effects of reagents used to solubilize the drugs. The metal complex-based drugs like cisplatin can be encapsulated into the core of micelles formed by block copolymers using PEG-poly (aspartic acid) (PEG-PAsp) or PEG-poly(glutamic acid) (PEG-Pglu) via a coordinate bond between platinum and carboxyl group in aqueous solutions absent of competitive ligands [50–54]. It was noticed that micelles prepared using PEG-PGlu exert much stronger anti-tumor activity due to increased hydrophobicity which results in higher degree of accumulation in tumor tissues compared to PEG-PAsp micelle core as the PGlu core has additional methylene group on each amino acid unit of the drug-loading block [51, 52]. A simple strategy can be used to design pH responsive polymer materials which can change their charge or hydrophilicity based on the environment. They show that spontaneous self-assembly with the change in pH can be used to deliver sensitive drugs [55, 56]. For example, Doxorubicin was delivered with an increased release rate when the pH is lowered from 7.4 to 5.5 [57]. Thus, they insure more efficient delivery of therapeutics in the target cells. It was further demonstrated that Dox-conjugate prepared using poly(ethylene glycol) (PEG) and a homopolymer poly(2-(diiso­ propylamino)ethyl methacrylate) PDPAEMA (which showed a transition from hydrophobic to hydrophilic) also showed pH-dependent response for the release of drug, i.e., at pH 7.4 only 10% of Dox was released while as the pH was lowered further to 5.5 over 90% of Dox was released in 36 h [58]. Polymeric micelle encapsulates the drug during the emulsification process and is released during the degeneration process due to either change in temperature or pH [59]. The micelles are easily degradable

Multifunctional Nanomedicine  373 structures so to increase their stability ionic gels were formulated by crosslinking the core with Ca2+ ion for encapsulating cisplatin and doxorubicin [60, 61]. Recently, various mixed micelle formulations prepared using aliphatic polycarbonates, containing urea and acid functionalized block copolymers are shown to have higher kinetic stability and thus accumulation in tumor is increased to greater extent than the formulations having lower kinetic stability [62]. A polymeric micelle encapsulated drug Placitaxel is a well-known nanoformulation used for metastatic breast cancer [63].

12.3.1.3 Proteins and Peptides for Drug Delivery Natural biomacromolecules, such as proteins and peptides, are attractive carriers for small drug molecules due to their inherent biodegradability, biocompatibility, low toxicity, and high aqueous solubility. The peptides and protein drugs prepared by recombinant technology produce replicas from natural sources. Peptides, proteins, and antibodies have been extensively used as ligands to selectively direct drugs. For proteins and peptides to be good therapeutic drug carrier, it should have good pharmacokinetic and pharmacodynamics properties. Peptides can be cell penetrating, cell targeting, and organ targeting. Cell penetrating peptide can transport drug through membrane translocation, whereas cell and organ targeting peptide act by interacting with receptor present on a particular cell [64]. These molecules target overexpressed receptors or antigens by particular cells or tissues. Ailments that might be treated more effectively with this class of therapeutics include autonomous diseases, cancer, mental dis­orders, hypertension, and certain cardiovascular and metabolic disease. One such receptor is integrin which is having a high binding affinity for Arg-Gly-Asp sequences, thus making it a preferable tool for targeting of drugs [65]. Peptides around less than 30 amino acid residues are used in cell penetrating mechanism either by direct penetration into the membrane or by endocytosis mediated entry. The SV40 peptide known as nuclear localization sequence can be used efficiently to deliver Pt(II). The N-terminal is modified using malonic acid derivative and carboplatin like complex is directly formed on the peptide from Pt(II), thereby resulting in increase in solubility of complexed carboplatin formed as compared to native carboplatin [66]. Similarly, FeroceneNLS conjugate and cobaltocenium–NLS conjugates are prepared which showed increased cellular uptake resulting in increased cytotoxicity [67, 68]. The stability in physiological buffers, the lack of toxicity and sensitivity to serum, and rapid delivery has made cell penetration strategy a

374  Advances in Metallodrugs potential method for clinical applications. Monoclonal antibodies are considered as promising therapeutic tool. The Fc receptors expressed on the surface of immune cells are capable of activating immune response on the antibodies. The antibody-coupled nanoparticles can be regarded as a very attractive drug-targeting system due to their advantageous ligand specificity and physical stability. Herceptin is a therapeutic agent which is also used in drug delivery systems. Monoclonal antibodies, when combined with a drug, can prove to be more beneficial than the drug alone or antibody alone in inhibiting the proliferation of cancer cells. They bind to the target thus signaling apoptosis, blocking MDR, and inhibit the DNA repair mechanism of target cells. Antibodies can also be used for specific cancer cells like cetuximab, an antibody used for pancreatic cancer cell lines which are having variable EGFR expression. They have been used to target nanoparticles to specific cancer types [20]. Albumin, casein, ferritin, and collagens have also been explored as drug delivery systems [69]. Protein-based NPs are small in size (up to 130 nm), biocompatible, and biodegradable with low toxicity. Albumin shows stability in variable pH range from 4 to 9 and temperature (4°C–60°C). Abraxane, albumin encapsulated paclitaxel has been approved by FDA for metastatic breast cancer as the first line of therapy. Paclitaxel and docetaxel-loaded albumin nanoparticles are currently in phase II of clinical trials. Deep penetration, good bio-distribution, and higher plasma clearance made albumin nanoparticle an ideal carrier for breast cancer [70].

12.3.2 Drug Delivery by Inorganic Nanomaterials Organic nanomaterials provide broad design flexibility for combining multiple functionalities, but this flexibility is combined with drawbacks including intrinsic design complexity, high manufacturing cost, and structural instability.

12.3.2.1 Metal and Metal Oxides Inorganic nanoparticles attract great interest in the field of nanomedicine delivery over bioorganic nanomaterials such as liposomes, dendrimers, peptides, and micelles due to tunable shape, size, unique optical, physical properties, and more control over surface properties [71, 72]. Metal nanoparticles are conjugated with the drugs of interest using different coatings such as dextrans, polystyrene, PEG, chitosan, and PEI [73].

Multifunctional Nanomedicine  375

12.3.2.2 Au NPs Gold nanoparticles have become an important tool in biomedical nanomedicine applications as they demonstrate excellent optical properties, exhibit surface plasmon resonance, good photothermal properties, show non-toxicity, and flexible surface chemistry. They act as good drug delivery agents when conjugated with smart materials which release the drug by applying some external stimulus [73, 74]. Gold nanoparticles are prepared in many shapes ranging from nanorods, nanospheres, and nanocages to nanoshells. They are readily attach to thiols, phosphines, and amines to alter their surface properties, thus allowing easy conjugation to S or N containing ligands such as citrates, peptides, lipids, and antibodies [75]. For example, PEG surface modified Au NPs were loaded with copper (II) diacetyl-bis(N4- methylthiosemicarbazone) complex used for targeted drug delivery [76]. Similarly, it was experimentally demonstrated that placitaxel conjugated Au-PEG-TNF injected intravenously showed greater degree of accumulation in colon carcinoma and reduced the tumor size [77, 78]. Gold NPs are shown to conjugate effectively with oligonucleotides (OEA-CD-NP), thus showing a successful delivery of DNA plasmids to breast cancer cells (MCF-7) [79]. The cytotoxicity of DOX against human glioma cell line LN-229 was shown to be enhanced with AuNPs loaded with porphyrin-capped containing DOX [80]. To increase the dispersability of drug in aqueous medium, gold nanoparticles were cross linked with thiol ended PEG and the anticancer drug was encapsulated in the delivery vehicles which was released on applying laser radiations [81]. Surface modified gold nanoparticles are suggested to be promising candidate for delivering therapeutics in case of brain tumor therapy due to their excellent conjugating capacity, low intrinsic toxicity, great stability, and ability to cross the BBB [82].

12.3.2.3 Carbon-Based NPs The unique physicochemical properties of carbon-based nanostructures are being explored for their multi therapeutic potential and their easy modification in a variety of structural formations, including carbon nanotubes, nano diamonds, carbon dots, graphene, and fullerenes have led to variety of applications in the field of nanomedicine. Among all of them, carbon nanotubes are reported to be used most for drug delivery. Carbon nanotubes (CNTs) are allotropes of carbon formed by rolling the graphene sheets which results in formation of single-walled carbon

376  Advances in Metallodrugs nanotubes (SWCNTs) and multiwalled carbon nanotubes (MWCNTs) having diameter varying from 1 nm to 20 nm. The CNTs are known for drug delivery due to high drug loading capacity as they possess large surface area. They can be used for imaging and detection [83], used for direct cytoplasmic delivery of conjugated molecules via “needlelike” penetration mechanism [84]. Biocompatibility of CNTs can be improved by modifying the surface properties by anchoring amphiphilic block polymers [85] or by dendrimers [86]. The drug can be immobilized by various methods on CNTs like either by chemical adsorption, encapsulation inside the CNTs [87] or by anchoring on the surface of functionalized CNTs [88, 89]. CNTs can be used as a carrier to amphotericin B which is a antimicrobial agent, thus reducing its antifungal toxicity in mammalian cells as compared to free drug [90, 91]. CNTs can be used in dentistry to enhance the stiffness of hydroxyapatite coatings which is the main substance of our teeth [92]. Cisplatin, a well-known anticancer drug, can be encapsulated in MWCNT and delivered at the target site without getting degraded [93]. In another report, chitosan functionalized SWCNT were attached with anticancer drug doxorubicin and it was concluded that lower dose of it is required than free DOX [94]. SWCNTs are also known for platelet activation, aggregation, and platelet granulocyte complex formation [95]. The enhanced drug loading capacity was observed in case of both SWCNT and MWCNT, to deliver the flat aromatic tetracyclic anticancer anthracycline drug DOX by attaching via π-π interactions which is a type of non-covalent interaction functionalized by PEG, thus enhancing the drug loading capacity and cytotoxicity for breast cancer cells. The release of DOX is further affected by the diameter of CNTs like they interact more strongly with larger diameter MWCNT as they possess bigger and flatter graphitic side walls that facilitate more efficient interaction [96, 97]. Dapsone, a drug possessing anti-inflammatory and antimicrobial activity used to treat malaria, pneumocystis pneumonia, leprosy, and dermatitis, was successfully delivered using MWCNTs which provided it better aqueous dispersability rapid ingestion without much toxicity [98, 99]. A xanthine derivative theophylline used for the treatment of respiratory diseases was delivered using surface modified CNTs/alginate microspheres. The CNT diminished the drug leakage, enhanced drug loading capacity, conferred greater mechanical strength, and sustained release of drug from the hybrid microspheres [100]. CNTs can also be used as vaccine delivery tool by acting as a template for bioactive peptides as in case of foot and mouth disease virus (FMDV), a B-cell epitope was covalently bonded to amine groups present on CNT

Multifunctional Nanomedicine  377 using a bifunctional linker. The peptides were recognized by specific monoclonal and polyclonal antibodies as they adopt a secondary structure around the CNT. Immunization with this conjugated peptide shows high antibody response as compared to free peptide. The protein responsible for recognizing, clearing, and killing pathogens in human immune system further confirmed that CNT can be used as novel vaccine delivery tool [101, 102]. In recent years, single layered surface modified graphene oxide is also used for drug delivery and gene delivery as efficient carrier due to its excellent physicochemical properties and mechanical strength. Graphene oxide can be used for delivery of cancer drugs after functionalization of its surface using PEG, folic acid, and other binding agents such as it can be used for loading of aromatic, insoluble SN38 drug [103], DOX [104], and camptothecin [105]. Nanodiamonds (ND) are nowadays investigated as platforms for localized drug delivery, targeted chemotherapeutics, and biocompatible imaging [106–109]. Nanodiamonds are effective in carrying wide range of water soluble and insoluble drugs while preserving their efficacy [110]. They are considered to the least toxic nanoparticles in carbon family [111] and inexpensive for mass production [109]. They were found to effectively deliver doxorubicin (DOX) and epirubicin [107, 112]. Carbon dots (CDs) are carbon-based nanomaterials which are discrete, quasispherical nanoparticles, with sizes below 10 nm [113]. CDs find wide application in drug delivery, bioimaging, sensing due to their fantastic features of biocompatibility, chemical inertness and solubility, cheap, and easy synthesis. Red emissive C-dots can be used in photcoustics and thermal thernostics for cancer diagnosis and treatment in biomedical field as can be seen from the data shown in Figure 12.4 [32]. A well-known anticancer drug DOX was loaded on CD using π-π interactions shows controlled drug release and excellent tumor therapy. Similarly, nitrogen and phosphorous dual doped CDs showed electrostatic hydrogen bonding interaction with DOX drug molecules [114]. DOX prolonged release and accumulation in the cells can further be enhanced using amine functionalized CDs conjugated via a linker, N hydroxysuccinimide ester-poly(ethylene glycol)10 maleimide resulting in higher tumor efficacy and less side effects as compared to free DOX [115–117]. Although CNT-based delivery system is quite successful, still longterm toxic studies need to be done before they reach to the level of clinical application.

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Figure 12.4  Photochemical and photophysical properties of C-dots. (a) The absorption of water-dispersible C-dots (100 µg ml−1). The insets show enlarged absorption of the C-dots from 600 to 750 nm. (b) Temperature elevation of pure water and the aqueous dispersion of C-dots with different concentrations under irradiation from a 671 nm laser with a power of 2 W cm−2 as a function of irradiation time (0–600 s). (c) Plot of temperature change over a period of 600 s versus the aqueous dispersion of C-dots with different concentrations. (d) Temperature IR images of C-dots aqueous solution under 671 nm laser (2 W cm−2) irradiation recorded with an IR camera at different concentrations, and (e) photothermal effect of C-dot aqueous solution (100 µg ml−1) when illuminated with a 671 nm laser (2 W cm−2). The laser was turned off after irradiation for 12 min. (f) Plot of cooling time versus negative natural logarithm of the temperature driving force obtained from the cooling stage as shown (e). The time constant for heat transfer of the system was determined to be τs = 345 s.

12.3.2.4 Silicon-Based Nanostructures for Drug Delivery Many inorganic nanomaterials, and even organic nanostructures, exhibit poor bio-compatibility and should be coated with biocompatible materials for use in biomedical applications [118] as well they may not be cleared by body inside or eliminated by renal excretion. This can be done using Si or black phosphorous nanoparticles which show good biocompatibility as well as biodegradability. Si degrades in body to form orthosilicic acid which is exist already in human body and can be excreted out through urine [119, 120]. Si QDs-based probes can be synthesized as shown by Figure 12.5 [120]. Similarly, black phosphorous gives final degradation products which are non-toxic to the human body [121]. Mesoporous silica nanoparticles (MSNs) are used in controlled drug delivery [122, 124, 125]. The MSNs are homogenous and have low polydispersity index and high surface area for loading the drugs. The high percentage of silanol groups on the surface can accommodate lot of organic

Multifunctional Nanomedicine  379 HF/HNO3 Acid Etching H

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254 nm UV light

O

Carboxyl terminated SiQD

Micelle encapsulated SiQDs

000)

2

Hydrophobic core Hydrophilic head

Methoxy terminated

e 00) Amin

(PEG 20 ) + DSPE

EG

DSPE (P

DSPE (PEG 2000) + DSPE (PEG 2000) Folate Funtionalized Silicon

DSPE (P

EG) + D

Amine terminated

SPE (PE

G 2000 Folate terminated ) Carbo xylic Aci d EDC + Transferrin Carboxyl Terminated

biomolecule conjugated Micelle

Figure 12.5  Synthesis and surface functionalization of Si quantum dots.

molecules via covalent or electrostatic interaction. MCM- 41 and SBA-15 are best known MSNs used for drug delivery [123]. The simulated drug profile of Ibuprofen loaded in MCM 41 with a pore size of 2.5 nm showed a release into the solution over a period of 3 days [124]. Varied range of drugs such as anticancer drugs [125], antibiotics [126], and heart disease [127] drugs can be loaded on silica nanoparticles either by physical or chemical adsorption [128]. A stimuli triggered superparamagnetic nanoparticle Fe3O4 capped MSN system was developed which can act a site selective sensory and for controlled release drug delivery [129]. Silicon quantum dots are considered good carriers of drugs into cells. Silicon quantum dots are known to deliver siRNA into tumor cells [130]. Besides this, encapsulated using poly(ethylene glycol)-block-polylactide (PEG-PLA), fluorescent Si quantum dots deliver the anticancer drug quercetin effectively to suppress human hepatoma HepG2cell proliferation [131]. However, silica particles have great potential for diagnostic and therapeutic applications but recent studies suggest both in vitro and in vivo toxicity of using nano silica. Silica nanoparticles are known to generate reactive oxygen species [132], induction of oxidase enzymes [133], and various other disturbances in cell, thus resulting in cell membrane damage.

12.3.3 Photo Therapy Photoinduced therapy is a non-invasive approach used for treating diseases such as cancer and other peripheral diseases using light. Two types

380  Advances in Metallodrugs of phototherapy are known, one is photodynamic therapy, in which photo­ sensitizer (PS) produce reactive singlet oxygen species to kill the disease affected cells [134–136], while the other one is photothermal therapy (PTT) that uses a photothermal agent to convert light energy to heat in the localized cell area to kill the affected cells [137, 138].

12.3.3.1 Photodynamic Therapy The basic principle in case of PDT is activation of a photosensitizer to induce a set of chain reactions by using light of a particular wavelength, thus generating cytotoxic reactive oxygen species (ROS) that can result in localized cell ablation [134, 139]. The induced chain reactions can take place by any of the two mechanisms, Type I or Type II. In case of Type I, the excited PS involves electron transfer resulting in formation of a free radical which then hamper the function of nucleic acid, fatty acids, and certain amino acid that lead to cell death [140]. The type II involves direct transfer of energy from excited PS to convert ground state triplet oxygen to excited state singlet oxygen and other reactive oxygen species, thereby creating an oxidative stress thus resulting in irreversible damage to mitochondria and other cell organelles and eventually selective cell death [137]. PDT treatment can cause cell death by any of the three mechanisms, apoptosis, necrosis, and autophagy. The cell death by apoptosis is most common one, followed by necrosis which occurs if type II mechanism is followed. Autophagy is the new mechanism proposed according to which the stress caused by the treatment cascades signals that leads to autophagy of cell. Nowadays, nanoparticles are used to conjugate or load photosensitizers for therapy. Nanoparticles have high surface to volume ratio which slow down the release of photosensitizer from the system into the blood stream. They also prevent degradation of the photosensitizer in the biological fluids thus improving the overall efficiency of therapy. It is observed that the currently used PS drugs like photofrin and several other clinically accepted second generation PS drugs like porphycenes and pthalocyanines have poor water solubility and selectivity. Thus, a combination of PS drugs with nanomaterials such as upconversion nanoparticles, quantum dots, carbon nanomaterials, and polymeric nanomaterials are used for both in vitro and in vivo PDT. For targeted delivery, peptides are conjugated to NPs such as ProtoporphyrinX (PpIX) conjugated with cyclic RGD peptide (sequence RGDFK) has shown good results in case of mouse carcinoma cells CanNT [141]. Similarly, PEGylated silicon pthalocyanine conjugated gold nanoparticles as well as ZnO nanorods conjugated with substituted prophyrins were developed for treating breast

Multifunctional Nanomedicine  381 cancer cells [142]. Several reports are known for destruction of cells by necrosis or apoptosis when semiconductor nanoparticles are used such as TiO2, ZnO, and Fe3O4 [143, 144]. Quantum dots are one more class which have found great usage in PDT due to their unique property of two photon absorption cross section along with high large volume to surface ratio. Biocompatible CdSe-porphyrin conjugate was synthesized which were water soluble and shows the production of singlet oxygen at high quantum yield known to have cytotoxic effect during PDT [145]. Recently, CdTe-Pthalocyanine conjugate is known to show high triplet quantum yield (0.90) [146]. Upconversion nanoparticles such as NaYF4 coated with mesoporous silica and loaded using ZnPc showed great efficiency in case of treating murine bladder cancer cells by producing singlet oxygen which can be monitored by fluorescent marker dye [147]. The photosensitizers get attached to the surface upconversion nanoparticles by covalent linkage, thus providing them superior stability and preventing their leakage during systemic circulation. Chitosan coated upconversion nanoparticles using ZnPc photosensitizer are known to improve photostability and dispersivity which enhances the production of singlet oxygen [148]. Gold nanoparticles are another class which has been used extensively by conjugating them with protein antibodies. As in case of targeted delivery where anti-HER2 monoclonal antibody was conjugated with gold nanoparticles when illuminated using HeNe laser results caused increased cell toxicity. Similarly, in vivo PDT of C57 mice bearing amelanotic melanoma shows damage to cells by disrupting the blood capillaries and endothelial cells [149]. Biodegradable polymers like chitosan and hyaluronic acid-based nanoparticles have received attention in their usage for both in vivo and in vitro PDT method due to their high drug loading capacity, biocompatibility, and non-toxic nature [150]. Chlorin e6-hyluronic acid nanoparticle conjugate injection in mice tumor cells was use to suppress the tumor by the singlet oxygen and intense fluorescence produced due to irradiation [151]. Thus, PDT is gaining more acceptance as a localized treatment method specifically for tumor cells but has not used as first line treatment method. It is a promising avenue if we can improve on the properties of photosensitizers, their degradability, loading capacity on nanoparticles, thus enhancing their selective accumulation resulting in enhanced cytotoxicity.

12.3.3.2 Photothermal Therapy Photothermal therapy (PTT) is a laser induced, non-invasive method of relying on an optical absorbing agent known as photosensitizer which can absorb energy and convert it into heat on stimulating by electromagnetic

382  Advances in Metallodrugs radiations, thus inducing programmed cell death in localized environment, thus reducing damage to healthy tissues [139, 152]. It uses near infrared (NIR) radiations which penetrated deeply inside the tissues with minimal scattering unlike PDT which cannot be used for solid tumors. Inorganic nanoparticles can convert energy from external sources into heat, which can be utilized for therapeutic purposes. When metal nanoparticles are illuminated, the valence electrons undergo a collective oscillation, thus leading to localized surface plasmon resonance (LSPR). This depends on shape, size, and type of metal from which nanoparticle is prepared [153, 154]. The dimensions of nanospheres used play a major role as with increase in size the LSPR shows red shift due to electromagnetic delay in larger particles. Huang et al. demonstrated that average temperature of cells should reach from 70°C–80°C to destroy cancerous cells in vitro [151]. Other way to tune the SPR is by using metal nanoshells like in case of silica NPs encapsulated in gold nanoshells, thus shifting the SPR towards NIR due to enhanced coupling between inner and outer surface plasmons of shell with decrease in shell thickness [156, 157]. This treatment was used to kill murine colon carcinoma cells using PEGylated gold nanoshells [158]. Photothermal ablation was successfully observed in a gold nanoshell conjugated monoclonal antibody, trastuzumab, for breast cancer cells [159]. Gold nanoparticles conjugated to pH sensitive ligands shift the absorption band to longer wavelength due to formation of gold aggregates with the change in pH [160]. This results in excitation of PT agent in vivo. PTT can be combined with drug delivery to enhance the efficiency as seen in case of doxorubicin encapsulated within PLGA nanoparticles with a gold layer deposited on the surface. Irradiation of the polymer conjugate degrades the PLGA layer thereby releasing the drug along with the PTT ablation of the cells thus resulting in therapeutic efficiency [161]. It has been shown that gold nanorods conjugated to antibodies like EGFR leads to cell death of malignant cells at lower energy than non-malignant cells due to presence of double the amount of nanorods being present on the cell surface [162]. Carbon nanomaterials are known to have high NIR absorption coefficient due to elongated conjugation band which makes them an ideal candidate for using in PTT method. Graphene an allotrope of carbon has covalently bonded carbon atoms into hexagonal packed 2D network which shows unique electronic and plasmonic properties which can convert energy of light to heat [163]. The reduced graphene oxide sheets show six times enhanced light absorption at 808 nm due to improved π conjugation in the reduced form maintaining its water solubility [164]. Recently, several graphene analogs such as carbides and sulfides are also used in PTT as they show excellent photothermal properties [165, 166]. Combined PTT

Multifunctional Nanomedicine  383 with PDT makes it more effective as in case of MoS2 nanosheets which show photothermally enhanced photodyanmic therapy using Ce6 as a PS agent whose delivery is enhanced after stimulating photothermally [167]. Similarly, PEGylated-BODIPY (boron dipyrromethene) showed simultaneous generation of singlet oxygen and heating of cells on introduction of iodine inside the BODIPY core and exposed to light source thus possessing excellent phototheraputic effects [168]. Multimodality is seen in using PEGylated graphene oxide (GO) conjugated with Fe3O4 for magnetic field assisted delivery of DOX, thus forming a GO-Fe3O4-PEG-DOX platform for selective damage of murine breast cancer 41 cells within external magnetic field [169]. Thus, combining PTT with other therapies such as immunotherapy could be promising as it can photothermally induce cancer cell death by the response of patients own immune system and potentially tumors outside the scope of laser radiation can also be treated.

12.3.4 Radiation Therapy Radiation therapy involves delivery of high intensity radiations including X-rays, Gamma rays, and high energy particles with high accuracy to tumor tissues resulting in death of tumor cells. This is one of the key treatments for cancer patients along with surgery and chemotherapy but its major disadvantage of this therapy is its non-specificity due to which significant number of healthy tissues also gets affected and acquired radiation resistance which leads to tumor relapse. So, nanoparticles can be used effectively to increase the radiosensitization of tumor cells and confining the radiation to affected cells. Radiotherapy works by damaging the DNA through production of water radicals and other reactive species, thus causing 70% damage and rest 30% due to secondary electrons produced and direct fragmentation of the DNA [170]. Using nanomaterials improves the radiation therapy by inducing more toxicity for tumors at low radiation level as the densely packed metals selectively scatter the radiations to generate photons, photoelectrons, Auger electrons, and Compton electrons [171]. Nanomaterials having high Z value such as Iodine are used for this purpose [172]. It was found that in treating brain tumors in mice radiotherapy plus Iodine nanoparticles combined with drug Doxil increased the lifetime by more than two fold compared to radiotherapy alone [173]. Nowadays, Gold nanoparticles are considered more promising as they have high Z than I2 and show more biocompatibility, are inert, show low clearance rate as compared to I2, and can be easily varied in size and shape, easy imaging, and quantification [174, 175]. AuNPs upto 50 nm size showed highest uptake by the cells [176]. Gold nanoparticles along with

384  Advances in Metallodrugs radiotherapy show multifold increase in the single and double stranded breaks in DNA of the cells when used along with other chemotherapeutic agents such as cisplatin [177]. Irradiation of 26 Gy and 250 kVp of X-rays for ∼1 min after the IV injection of 2.7 g Au kg−1 resulted in 86% long-term survival (>1 year) compared to identical irradiation without gold which showed only 20% survival [178]. Cisplatin binding to guanine was shown to result in better bond dissociation by triggering the formation of transient anions. Gadolinium is another metal used as radiosensitizer as they can generate long lived pi-radical cations upon exposure to hydrated electrons. Its in vivo effectiveness was studied in case of murine carcinoma cells [179]. Superparamagnetic Iron nanoparticles also known to enhance the radiation induced DNA damage by catalyzing the reactive oxygen species, thereby creating an oxidative stress inside the cell [180]. Hafnium oxide nanoparticles (NBTXR3) were also being evaluated as a radiosensitizer for colorectal cancer cell line where it showed delayed tumor growth [181]. It is in phase I clinical trials for soft tissue sarcoma (NCT01433068) and advanced squamous cell carcinoma (NCT01946867). It is concluded that adding nanoparticles to the tumor can enhance both the dose and contrast at the target in radiotherapy.

12.3.5 Neutron Capture Therapy Neutron capture therapy is used for gliomas specifically in brain which are resistant to all current forms of therapies known including surgery, chemotherapy, and radiotherapy to name some. These are basically malignant cells which infiltrate beyond the margins of resection and both chemo and radiotherapy dosage is compromised due to normal tissue tolerance limit [182, 183]. To successfully treat these micro-invasive tumor cells, a therapy must be developed to selectively treat the malignant cells with no or little effect on normal cells. For this, certain elements can be used which can absorb slow moving or thermal neutrons which have the potential of having their radiant energy confined to the cell from which they arise and it is of lethal magnitude [184]. Boron neutron capture therapy (BNCT) is one of this type where non-radioactive boron-10 is irradiated with low energy thermal neutrons, thus resulting in fission reaction 10B(n,α) 7 Li-capture which results in destruction of cells containing sufficient number of 10B atoms [185–189]. Boron compounds which are used as potential 2 2 , B12H12 candidate for BNCT are sodium decahydrodecaborate, B10H10 anions, carboranes, and their derivatives [184, 190–192]. (L)-4-dihydroxyborylphenylalanine, referred to as boronophenylalanine or BPA, is one of the low molecular weight boron compound which shows promising results

Multifunctional Nanomedicine  385 by persisting for long hours in animal tumors [193]. Boron clusters can also be used to treat arthritis [184]. Boron clusters may be conjugated to growth factors such as EGF (epidermal growth factor) [194, 195]. Thus, neutron capture therapy can be used as a successful therapy by developing compounds which can achieve specific tissue targeting at a concentration level with acceptable degree of systemic toxicity.

12.4 Regenerative Medicine It involves restoration of tissues which have lost their functionality due to physiological reasons or genetic reasons. The main aim is basically constructing scaffolds that activate regeneration of tissues [196]. The synthetic constructs used as scaffolds should promote the migration and development of cells to form tissues and should be biodegradable and biocompatible [197]. The compatibility factor can be taken care by using host organism’s cells or proteins to create the scaffolds. Latest research shows that stem cells, undifferentiated ones can also be used and can be converted into almost any desired cell type by cell culture [198, 199]. Gene therapy can also be used which involves delivery of missing or defective proteins, growth promoters, or other immunological components. It basically deals with restoration of functional tissue when the patient body’s own response is not sufficient to regenerate healthy cells. These include regeneration of bones, cartilage, tendons, and skin, promoting wound healing in case of soft tissues and burns and in patient suffering from diabetes mellitus and neurogenerative disease [200]. World over various research institutes are working in this field taking help of nanotechnology to heal and treat the tissue damage at source rather than their symptoms, thereby enhancing the quality of life. Nanomaterials can match the nanoscale biological cells and thus can play important role in generating materials for tissue regeneration. Thus, nanotechnology has become more relevant in case of tissue engineering and manufacturing of biomaterial scaffolds which closely replicate the structure of extra cellular matrix. A wide range of nanomaterials used for tissue engineering are carbon nanotubes, nanofibers, nanowires, and nanospheres which serve various applications such as acting as biosensors, as scaffolds for tissue growth, delivery devices for various genes, and growth factors. To provide biomimetic environment, natural materials can be used as scaffolds like collagen, fibrin which provide better cell adhesion and proliferation, migration, and extracellular matrix deposition. But these natural materials suffer from various drawbacks such as immunogenicity, can be easily attacked by pathogens, less

386  Advances in Metallodrugs mechanical strength, and easy degradation [201, 202]. Thus, lot of synthetic nano scaffolds are used nowadays as in case of cartilage tissue engineering nano amorphous calcium phosphate with poly (L-Lactic acid) is used to repair cartilage defects in rabbits. It was observed that when the material was implanted with basic fibroblast growth factor, most of the defects are filled [203]. Superparamagnetic nanoparticles are also used for developing magnetic scaffolds for osteochondral defects which provides a unique possibility to adjust scaffold activity according to patient’s personal needs such as accumulation of growth factors and drug doses [204, 205]. Nanostructured beta tricalcium phosphate coatings on scaffolds enhance biological properties [206]. Hydroxyapatite has a strong tendency to attract the osteoblasts so it can be combined with various natural and synthetic polymers including PLA (poly(L-Lactic acid), PLGA, collagen, chitosan forming composite scaffolds having improved porosity, degradability, and mechanical strength [207–209]. It was also observed that skin regeneration in case of diabetic animal using scaffolds of copolymer PCL/poly-ethylene glycol and chemically modified using human epidermal growth factor showed improved degree of wound healing [210]. Thus, these tissue engineering can be used at clinical level for cartilage regeneration [211], nerve regeneration [212], myocardial [213], ocular [214], and dental regeneration [215]. Integration of nanomaterials into scaffolds no doubt enhances biological regulation of cell behavior for regeneration but detailed study regarding this technology to be used at commercial level is underway which can give a clear insight about its the clinical efficiency, cost effectiveness, and cytotoxicity of nanomaterials used before making it a hopeful solution.

12.5 Future Prospects and Conclusion Nanomedicine is considered a disruptive technology which opens a wide array of possibilities for human control over health by increase in its efficiency and precision. Usage of nanomedicine has introduced new methods of diagnosis and improved the efficacy of treatment whether in drug delivery, various medical therapies, tissue engineering, thus taking the medical field to another level of automation and personalized medication. But along with this, we also should try to understand in more detail the mechanism of toxicity of the currently used nanomaterials in various applications thereby developing new materials with low toxicity and expanding the medical application of nanomaterials for human health.

Multifunctional Nanomedicine  387

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Index 2D-DIGE, 260 5 Phosphodiesterase (PDE5), 180 56MESS, 222, 228 Acetaldehyde, 162 Acetylacetonate, 20 Acetylated, 326 Acridine, 218 Adenoviral conjunctivitis, 16 African sleeping sickness disease, 15 AGU conformatiom, 340 Ajoenes, 183 Alkyl nitrites, 166 Alopecia, 8 Aluminum hydroxide, 17 Aluminum phosphate, 17 Amorphous cellulose, 338 Amsacrine, 219 Amyl nitrite, 165 Angina pectoris, 166 Angiogenesis, 206 Angiogenesis as a substantial target of tumorigenesis, 300 Animalic spirit, 162 Anthracene, 218 Anthraquinone, 218 Anti-angiogenic, 256 Anti-apoptotic, 159 Anticancer activity, 44 Anticancer activity for breast cancer, 183 Anticancer compound, definition, 206–207 Anticancer copper(II) complexes, 82

essential for health and nutrition, 82–83 Anticancer drugs, 167 Anticancer metallodrugs, 6–14 Anticancer platinum(II) complexes, 24 Anti-inflammatory, 159 Anti-inflammatory properties, 175 Anti-leukemia activity, 166 Antimalarial drug, 16 Antimicrobial activity, 139 Antimicrobial metallodrugs, 15–16 Antimony-based complexes, 15 Antineoplastic effects, 12 Antioxidant behavior of H2S, 179 Antioxidant features of garlic, 183 Anti-proliferative, 159 Antitumor, 248, 250, 252–254, 256, 259 Antiviral metallodrugs, 16–17 Apoptosis in cancer cells, 171 Argyropoulos, 322 Arsenical, 2 Arsobal, 15 Ascorbic acid, 167 Atherosclerosis, 182 Atherosclerotic changes, 184 Auranofin, 4, 251, 252, 254, 268 Azidation, 331 B3LYP, 172 BBR3564, 217 Binding mode, 279, 281–283 Biological membranes, 166

403

404  Index Biological importance of zinc, 90–92 Bioregulatory conditions, 168 Bleomycin, 7, 10 Blood pressure control, 164 Boric acid, 176 Breast cancer, 183 Broccoli, 184 Bromoisobutyryl bromide, 328 Butyl nitrite, 165 C. glycidyltrimethyl potassium, 336 C. albicans, 140 Cancer, 43 cells, 172, 205–206 types, 204–205 Cancer cell lines, 182 Carbon monoxide, 158, 162 Carbonmonoxy-myoglobin, 175 Carboplatin, 8–9 Carboranes, 384 Carboxy-hemoglobin, 173, 174 Cardiolite, 18 Carrageenan-induced knee joint synovitis, 178 Casiopeinas®, 11–12 Caspase-dependent apoptosis, 56 Catalytic metallodrugs, 22–23 Cellulose, 319 CH1, 255 Chalcones, 226 Chalcoplatin, 227 Chemotherapeutic, 249 Chemotherapy, 206, 207 Chemotypes, HID, 284–285 HNO, 284 HPD, 278, 280, 282, 284–285 Chloro(triethylphosphine)gold(I), 252 Chloroquine, 16 Cholesterol, 182 Chromium, 20

Circumstance, 325 Cis-tetrachlorodiamine platinum (IV), 250 Cisplatin, 7, 8, 50, 247, 248, 250, 251, 253, 255, 256, 259, 261, 263, 264, 265 Cisplatin antitumor activity, see also Pt complexes, 209, 215–216 Cobalt complexes, 138 CO-based drugs, 185 Collagen synthesis, 167 Colorectal carcinoma, 153 Combination therapy, 63, 225–226 Coordination complexes, 137 Copper, 11 Copper and human health disorders, 83–84 Menkes’ Disease (MD), 85 Wilson’s Disease (WD), 84–85 Copper complexes as potential therapeutic agents, 85–86 naphthoquinones, 88–89 quinolones-based copper complexes, 88 thiosemicarbazones-based complexes, 86–87 Copper(I)-catalyzed azide-alkyne cycloaddition (CuAAC), 282 CORM-A1, 176 CORMs, 174–176 Coronary arteries, 164 Coronary vasoconstriction, 175 Cruciferous vegetables, 184 CT-DNA, 151 Cutaneous inflammation, 169 Cyclophosphamide, 7 Cystathionine-β-synthase, 159 Cystathionine-γ-lyase, 159 Cytochrome c-oxidase, 165 Cytochrome P450, 174 Cytoplasmic blebbing, 143 Cytosolic GSTs, 167 Cytotoxic, 53

Index  405 Cytotoxic peroxynitrite, 171 Cytotoxicity, 158 Darinaparsin, 4 Daunomycin, 219 DEA-NO, 166 DeCyder, 261 DEDD motif, 274–276 Denitrification, 160 Deoxymyoglobin, 175 DFT, 172 Diabetes mellitus, 19 Diagnostics and imaging, 366–369 MRI, 366 OAT, 367 SPIONs, 367 Diallyl disulfide, 183 Diallyl thiosulfinate, 183 Diallyl trisulfide, 183 Diazeniumdiolates, 166 Dicarboxylate esters, 20 Diclofenac, 182 Diketo acid (DKA), 280–281 Distamycin, 214 Dithiocarbamates, 254, 262 DNA, 383 DNICs, 170 Dotarem, 19 Doxorubicin, 7, 219 Doxovir (CTC-96), 16 Dye-nitrosyl conjugates, 171 Dyson, P. J., 14 Edema formation, 182 Ellipticine, 218 Encapsulated, 372, 374–376, 379, 382 Endogenous production of CO, 158 Endothall, 227 Endothelial cells, 163 Endothelium-derived relaxing factor, 162

EPR, 363, 366, 368 Erectile, 181 Erectile dysfunction, 182 Escherichiacoli, 212–213, 250 ESI-Ion trap MS/MS, 261 Etoposide, 7 Exogenous NO, CO and H2S donors, 170 Extra cellular matrix (ECM) proteins, 206 Ferid Murad, 158 Ferrocene, 16 Ferrocene-based antimalarial agents, 117 mechanism of action, 118 Ferrocene-based antibacterial and antifungal drugs, 118 Ferrocenyl Guanidines as antibacterial and antifungal agents, 21 Schiff base derived ferrocene conjugates as antibacterial agents, 19 Sedaxicene as antifungal agents, 122 Ferrocene-based anti-tumor and anti-cancerous drugs, 123 Ferricenium salts as anti-tumor agents, 124 Ferrocene conjugated to peptides for lung cancer, 125 Ferrocenyl derivatives of retinoids potential anti-tumor drug, 27 Ferrocenyl sub-ordinates of illudin-M, 126 Ferrocenylalkyl nucleobases potential anti-cancerous drugs, 26 Ferrocenylalkylazoles active anti-tumor drugs, 124 Targeting breast cancer with selective ferrocene based estrogen receptor modulators (SERM), 28

406  Index Ferrochloroquine (antimalarial agent), 15 Ferrocifen, 11 Ferroquine, 16 Filamentous, 250 Flamazine®, 15 Fluorescent quantum dots, 368 Fluoroquinolone, 252, 268 Folate receptor (FR) proteins, 230 Formyl-Met-Leu-Phe-OH, 178 Fungal, 182 Gadovist, 19 Ganglion cells, 181 Garlic (Allium sativum), 182 Gaseous molecule, H2S, 157, 159 Gaseous signaling molecules, 161, 162 Gasotransmitter, 160 Gastric lesion, 180 Gastrointestinal effects, 103 Gastrointestinal ulceration and bleeding, 182 Gaussian 09W software, 172 Gene therapy, 385 Glaucoma management, 181 Glia, 161 GLOBOCAN, 248 Glucose transporter (GLUT), 21 Glutathione, 212 Glutathione pool, 255 Glutathione reductase, 170 Glutathione-S-transferase, 174 Glyceryl dinitrates (GDN), 163 Glycogen for (1–4)-glycosides bonds, 21 GSH, GSNO, GST, 166 GTMAC hydrolysis, 337 Guanylate cyclase, 159, 165 Gycerin trinitrate (GTN), 163, 170 GYY4137, 178 H2S, 157, 176, 177, 183 Healthcare applications of copper, 83

Heart failure and hypertension, 169 Heme oxygenase (HO), 159, 185 Hemodialysis, 348 Hemoglobin (Hb), 158 Hemorrhagic shock, 166 Hepatitis C virus (HCV), 23, 282 Heptaplatin, 8–9 Hexahistidine (H6), 274 HIV integrase, 278, 280 HL-60, 254 Hoechst 33258, 217 Horseradish, 184 HS-NSAIDs, 182 HT-29, 256, 259 Human cancer cells, 172 Hydroxyapatite, 386 Hypertensive effect, 168 Hypoxic cells, 62 Iberin, 185 IC50 values, 45 Ifosfamide, 7 IK-1001, 177 Imidazolium, 12 Iminophosphorane-organo gold(III), 260 Implementation, 334 Indazolium, 12 Influential hydroxyl, 343 Inhalation, 174 Inroganic nanomaterials, 374–379 carbon NPs, 375–377 gold nanoparticles, 375 silicon NPs, 378–379 Insulin mimetic metallodrugs, 19 Insulin receptor tyrosine kinase (IRTK), 20 Interaction of cisplatin species, 80–82 Intraocular pressure (IOP), 181 Iproplatin, 224 Iron, 10 Ischemia, 170

Index  407 Ischemia damage, 181 Ischemia reperfusion injury, 185 Isoamyl nitrite (IAMN), 165 Isobutyl nitrite (ISBN), 165 Isosorbide 5-mononitrate (ISMO), 163 Isosorbide dinitrate (ISDN), 163 Isothiocyanate compound, 184 Kidney, 175 Knee joint synovitis, 178 Kojic acid, 20 KP1019, 12–14 Latanoprost, 181 Later azidation, 341 Lawesson’s reagent, 178 L-cysteine, 170 LED device, 172 Leukemia cancer cell lines, 182 Leukemia L1210, 250 Levofloxacin, 252 Liposomal drug delivery, 369–371 Lisinopril, 22 Long terminal repeats (LTRs), 274 Lung disease, 174 Magnevist, 19 MALDI-TOF MS, 261 Malignancies, 248 Maltol, 20 Maltose-binding protein (MBP), 275 Mammalian tissues, 159, 163 Manufacturing, 342 MCF-7, 254, 256, 259, 262 MCF-7 cells, 185 Medicinal inorganic chemistry, 42 Melarsoprol, 15, 16 Melglumine antimoniate (Glucantim or Glucantime), 15 Melphalan, 7 Membrane receptor, 161 Messenger molecule, 162

Metal carbonyls, 176 Metal chelates, 43 Metal nitrosyl compounds, 168 Metal salts and their compounds, use of, 3–4 Metal-based antitumor chemotherapeutics, 7 Metallic nanoparticles, 364 Metallodrugs in anticancer therapy, 297 Metallodrugs, therapeutic, 6–26 anticancer metallodrugs, 6–14 antidiabetic metallodrugs, 19–21 antimicrobial and antiviral metallodrugs, 15–17 catalytic metallodrugs, 22–23 future, 23–25 radiopharmaceuticals and radiodiagnostic metallodrugs, 17–19 Metallopharmacuticals, 157 Metals and metallodrugs in angiogenesis, 302 Microsomal, 166 Mitoxantrone, 219 Modification, 333 Molecule of the year, 158 MRI, 18–19 MS-325, 4f Multifunctional nanomedicine, 363, 365 Mustards, 7 Myelosuppression, 8 Myocardial contraction, 178 Myoglobin, 176 Myoview, 18 NAMI–A, 12 Nanodrugs, 363, 371 Nanoparticle-based drug delivery systems, 25 Naphthalimide, 256, 269 NCI-H460 lung cancer, 220 Nedaplatin, 8–9

408  Index Nephrotoxicity, 8 NER inhibitor, 225 Neurotoxicity, 8 Neurotransmission, 158 Neurotransmitters, 161 Neutron capture therapy, 384–385 Nitrate-silver sulphadiazine (Flammacerium), 15 Nitric Oxide (NO), 157 Nitrogen donors of DNA strand, 80 Nitroprusside (SNP), 163 Nobel prize for medicine and physiology, 158 NO-donors, 167 NONOates, 166 Nonsteroid anti-inflammatory drugs (NSAIDs), 182 NO-releasing composites, 172 Norfloxacin, 252 Nucleophilic displacement reactions, 80 Oestrogen, 229 Oestrogen receptors (ER), 228 Ophthalmic instrument, 351 Organic nitrates, 163 Organic nitrites, 165 Ototoxicity, 8, 250 Ovarian, 250, 253, 255–257, 259, 260, 263, 270 Oxaliplatin, 8–9, 228–229 Oxidation, 324 Pacemaker, 167 Parasitic infections, 182–183 Patelet aggregation, 158, 170 Pathogenesis of human diseases, 162 Pathogens, 160 Pathophysiological functions, 162 Pentaerythrityl tetranitrate (PETN), 163 Pharmacological actions, 174 Pharmacologists, 185 Phenanthridine, 218

Phenanthroline, 218 Phosphodiesterase (PDE), 21 Photodynamic behavior, 172 Photodynamic therapy, 146 , 231, 380–381 Photo-excitation, 175 Photo-NORMs, 171 Photosensitive, 169 Photothermal therapy, 381–383 Physiology, 158 Picolinic acid, 20 Plant derived compounds, 182 Platinol, 137 Platinum anticancer drugs, 74–80 Platinum electrodes, 250 Platinum-based anticancer agents, 9 Pneuma, 162 Polyhydric alcohols, 163 Polymeric drug delivery, 371–373 Potassium alum, 17 Precursor decided, 342 Preeclamptic women, 164 Privileged substructure-based diversityoriented synthesis (pDOS), 282 Problematic meyhodology, 352 Prodrugs, 6 Pro-drugs, 174 Prostate specific membrane antigen (PSMA), 230 Proteasome, 251 Protein drug delivery, 373–374 Pt complexes, anticancer attributes of, 207–208 native state behavior, 208–209 Pt complexes as anticancer drugs DNA-coordinating Pt(II) complexes, 214, 217–218 DNA-covalently binding Pt(II) complexes, 219–222 photodynamic killing of cancer cell by, 231 Pt(IV) prodrugs, 224–230 multiple action, 25–228

Index  409 targeted, 28–230 targeted complexes, 222–224 Pt compounds in cancer treatment DNA adducts, 210–214 DNA as primary target, 209–210 DNA binding activities, kinetics, 210 Pthalocyanine, 380, 381 Pyrazole, 149 Radiation therapy, 383–384 Radiopharmaceuticals and radiodiagnostic metallodrugs, 17–19 Rearrangement exposures, 330 Regenerative medicine, 385–386 Resulting density, 332 Reverse transcriptases (RTs), 274 Rhenium carbonyl, 175 Rheumatoid, 251 RNase H active site, 274 RNase H function, 274–275 Role of zinc in human body, 97–98 Roussin’s red and black salt, 170 Ruthenium, 12–14 Ruthenium nitrosyls, 171 Ruthenium-based antitumor drugs, 43 Saccharomyces cerevisiae, 222 Salvarsan, 2, 15 Sarcoma 180, 250 S-aspirin, 179 Satraplatin, 8 Scaffolds, 344 Schiff base chemistry, 92 metal complexes, 92 hydrazone-type ligands, 3 Salen-type ligand, 3 thiosemicarbazone/ semi-carbazone ligands, 3 Selective targeting of CO release, 174 Signaling and regulatory functions, 157

Signaling molecules, 160 Silent killer, 158 Silvadene®, 15 Silver complexes, 15 Silver sulphadiazine, 15, 16 Single-walled carbon nanotubes (SWNTs), 230 SK-OV-3, 253, 263 S-Latanoprost, 181 Smooth muscle cells (SMC), 164 S-Nitroso-N-acetyl-pencillamine (SNAP), 166 Sodium hydrosulfide (NaHS), 177 Sodium molybdate, 21 Sodium nitroprusside, 168 Sodium stibogluconate (Pentostam), 15 Sodium tungstate, 21 Sodium–2–ethylmercurithio– benzoate, 17 Sparfloxacin, 252 S-Sildenafil, 180 Stents, 167 Steric considerations, 339 Strategies in drug discovery, click chemistry, 279 drug repurposing, 284 dual-target inhibitors, 280–281 high-throughput screening (HTS), 276 natural extracts, 283–284 pharmacophore model, 278 privileged substructure-based library, 282–283 Structure-activity relationships (SAR), 277, 285 Sulforaphane, 184 Sulphydryl (–SH) compounds, 15 SunPla, 9 Systemic toxicity of cisplatin, 82 Tamoxifen, 11 TDI modification, 335 Technetium–99m (99mTc), 17

410  Index Tert-butyl nitrite (TBN), 165 Testicular, 250 Therapeutic, 157, 160, 162, 248, 253, 264, 265 Thermoplastics, 320 Thioredoxin reductase, 254, 255, 257, 259, 268 Titanium anticancer agents, 10f Titanocenedichloride, 10 Top food sources of zinc, 95–97 Topical applications, 167 Topoisomerase II inhibitors, 7 Transesterification, 327 Triethylamine, 329 Trypanosomiasis (sleeping sickness) diseases, 15 Tumor suppression, 158 Ubiquilin-1, 261 Urinary system, 180–181 Vanadium-phosphatase protein structures, 20 Vanillic acid, 323 Vascular permeability, 158 Vascular smooth muscle, 166 Vascular stents, 167 Vascular tissue, 165

Vasculature, 165 Vasoconstrictors, 165 Vasodilator therapy, 166 Vasorelaxant, 162 Vesicles, 161 Viral, 182 Visible light irradiation, 172 Vitamin B12, 138 Vitamin E, 165 VT1161 (antifungal agent), 15 Wallflowers, 184 Warburg effect, 230 Wasabi, 184 Xanthine oxidase (XO), 166 Ylideneamine, 252, 268 Zinc, 21 Zinc anticancer complexes, 89–90 Zinc as a health benefit, 98–99 Zinc in alloy and composites, 100 Zinc supplementation as a treatment, 100–101 zinc and viral infections, 103 zinc deficiency, 101–102 zinc toxicity, 102 Zinc-based complexes, 93–94